CN103403557A - Microfluidic processing of target species in ferrofluids - Google Patents
Microfluidic processing of target species in ferrofluids Download PDFInfo
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- CN103403557A CN103403557A CN2011800631253A CN201180063125A CN103403557A CN 103403557 A CN103403557 A CN 103403557A CN 2011800631253 A CN2011800631253 A CN 2011800631253A CN 201180063125 A CN201180063125 A CN 201180063125A CN 103403557 A CN103403557 A CN 103403557A
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- G01N27/416—Systems
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Abstract
Disclosed are systems, devices, methods, and other implementations, including a device to detect at least one target species in a sample, with the device including a microfluidic channel configured to receive the sample containing the at least one target species and a biocompatible ferrofluid in which the at least one target species is suspended, a detector to determine the at least one target species in the sample, and at least two of electrodes positioned proximate the microfluidic channel, the at least two electrodes configured to generate controllable magnetic forces in the sample containing the ferrofluid when a controllable at least one electrical current is applied to the at least two electrodes. The generated controllable magnetic forces cause the at least one target species to be directed towards the detector. Also disclosed are devices for separating target species in a ferrofluid, and for focusing target species suspended in a ferrofluid.
Description
The cross reference of related application
The application requires on Dec 7th, 2010 to submit to, PCT application number PCT/US10/59270, exercise question is the U.S. Provisional Application sequence number 61/267 of " n cell that is undertaken by biocompatible ferrofluid operates and sorting " and submission on Dec 7th, 2009, the U.S. Provisional Application sequence number 61/407 that on October 28th, 163 and 2010 submitted to, 738 right of priority, its all the elements this by reference integral body incorporate into.
Field
The disclosure relates generally to the miniflow of ferrofluid and processes, and comprises the ferrofluid that comprises plurality of target material (for example biological target substance).More particularly, the disclosure relates to device, system and method,, to carry out these operations,, as separate the plurality of target material in biocompatible ferrofluid, concentrates target substance in biocompatible ferrofluid, detects target substance etc. in sample.
Background
The early diagnosis that relates to the disease (as metastatic carcinoma or low-level bacteremia) of rare cell in blood need to separate fast and accurately with the accurate monitoring of some genetic disease state (as sickle cell anemia), sorting and guide target cell type be towards sensor surface.In this respect, cell manipulation, separate and be sorted in cancer diagnosis (Dittrich etc., 2006,
Nat Rev Drug Discovery5:210-218), pathogen detection (Beyor etc., 2008,
Blamed Microdevices10:909-917) and genomic testing (Kamei etc. 2005,
Biotned Microdevices7:147-152; Cheong etc., 2008,
Lab Chip8:810-813) find just more and more application potential in the various biologicall tests under the background.
Have various noncontact microoperation methods, comprise the light tweezer (Ashkin etc., 1987,
Nature330:769-771; Chian et al., 2005,
Nature436:370-372), dielectrophoresis (DEP) (Hughes, 2002,
Electrophoresis23:2569-2582), based on the separation vessel of magnetic bead (Lee et al., 2001,
Appl Phys Lett79:3308-3310; Yan et al., 2004,
Phys Rev E70:011905) and determinacy fluid dynamics (Davis et al., 2006,
Proc Natl Acad Sci USA103:14779-14784).But a lot of existing methods can not realize fast reliably, high flux and resolution, simultaneously low-cost (Dufresne et al., 1998,
Rev Sci Instrum69:1974-1977; Kremser et al., 2004,
Electrophoresis25:2282-2291; Cabrera et al., 2001,
Electrophoresis22:355-362).The light tweezer provides operating single celled high resolving power and sensitivity, although these operations may cause sample heating (Liu et al., 1995,
Biophys J68:2137-2144), and usually be subject to minimum zone (Ashkin et al., 1987,
Science235:1517-1520).Holographic scheme light tweezer scope has been extended to recently touch simultaneously tens cells (Applegate et al., 2004,
Optical Express12:4390-4398), although total flux is still very low.
Based on the scheme of electric field, DEP for example, provide realize comprehensive, cost effectively device be used for operating simultaneously the potential of various kinds of cell.Yet, depend on to their performance sensitive concrete liquid medium electrical property, particle shape and its effective dielectric constant (Pethig et al., 1997,
Trends Biotechnol15:426-432).DEP device operation scheme needs careful optimization with the ionic medium of working for each different cell type, to reach in minimizing, heat (Menachery et al., 2005,
NanoBiotechnology152:145149; Muller, et al., 2003,
IEEE Eng Biol Med Mag22:51-61) and with polarization minimize (Sebastian et al., 2006,
J Micromech MicroengSpendable balance between needs 16:1769-1777).With the magnetic bead separate targets molecule of functionalization and cell by non-electric field overcomes these challenges with magnetic field.But the unfavorable aspect of this technology is long incubation time and clean cycle, and the difficulty of removal posteriority mark (Gijs 2004,
Microfluidics Nanofluidics1:22-40).By Davis etc. (Davis et al., 2006,
Proc Natl Acad Sci USAThe determinacy hydrodynamic method of 103:14779-14784) showing can be realized high-resolution separation, and does not use any electromagnetic field.But the high flux of this device need to high-resolution photoetching on large tracts of land, makes each installation cost keep high.
In biological medicine, the most common application of ferrofluid relates to the soliquid of the high dilution of magnetic nanoparticle.Its most widely commercial use be the MRI contrast preparation (Kim et al., 2005,
J Magn Magn Mater289:328-330).When suitably using the target antibody dressing, they can also be used for cancer hyperthermia therapy or as sensor detect pathogen (Scherer et al., 2005,
Brazilian J Phys45:718-727).
Although these development during ferrofluid uses provide much chances in medicine and diagnosis, this area still needs biocompatible ferrofluid is used for the controlled operation and the miniflow platform that separates fast of particulate and living cells.
General introduction
The device that is separated in the particulate samples that suspends in biocompatible ferrofluid has been described.In some embodiments, device comprises microchannel, and it has the length between sample inlet, at least one outlet and sample inlet and at least one outlet, and wherein sample can be added to sample inlet, and along length, flows at least one outlet.Device comprises a plurality of electrodes, and wherein microchannel length is crossed a plurality of electrodes, and further comprises the power supply that applies electric current to a plurality of electrodes, with the length along microchannel, produces magnetic field model.In some embodiments, interelectrode interval increases gradually.In some embodiments, reduce gradually at interelectrode interval.In some embodiments, a plurality of electrodes comprise at least one electrode layer.In some embodiments, a plurality of electrodes comprise a plurality of electrode layers.In some embodiments, a plurality of electrode layers are right angle patterns basically.In some embodiments, a plurality of electrodes comprise the concentric circles pattern.In some embodiments, the wall of microchannel length comprises zone small-sized, carinate, reeded, fluted or that tilt.In some embodiments, microchannel length with about 0-360(and more particularly, the 0-90 degree) between the angle at least a portion of crossing a plurality of electrodes.In some embodiments, particle is living cells.
The system of separating at least one target in the sample from be suspended in biocompatible ferrofluid has also been described.System comprises microchannel, and it has the length between sample inlet, at least one outlet and sample inlet and at least one outlet, and wherein sample can be added to sample inlet, and along length, flows at least one outlet.System also comprises a plurality of electrodes, and wherein microchannel length is crossed a plurality of electrodes, and when electric current is applied to electrode, further produces the magnetic field model along microchannel length.System further comprises at least one target in the sample that is suspended in biocompatible ferrofluid, and wherein when at least one target passed through along at least a portion of microchannel length, at least one target separated from remaining sample.In one embodiment, biocompatible ferrofluid comprises appropriate ionic species and promotes the cell continuation with the osmotic pressure of controlling on cell.In some embodiments, biocompatible ferrofluid comprises the approximately citric acid salt concentration of 5-200mM.In some embodiments, biocompatible ferrofluid comprises the approximately citric acid salt concentration of 40mM.In some embodiments, biocompatible ferrofluid has approximately 7.4 pH.In some embodiments, at least one target based target size separation.In some embodiments, at least one target based target shape is separated.In some embodiments, at least one target based target resilient separation.In some embodiments, target separates by being directed to selected outlet.In some embodiments, target is hunted down based on electrode gap.In some embodiments, at least one target is cell.In some embodiments, at least one target is particle.
The method of separating at least one cell type has also been described.Method comprises the following steps, two or more cell types are suspended in biocompatible ferrofluid to form sample, make sample by crossing the microchannel of a plurality of electrodes, apply electric current along microchannel length, to produce magnetic field model to a plurality of electrodes, and based on cell size, shape and flexible at least a difference sorting cells, enter at least one exit passageway.In one embodiment, cell separation efficiency is at least 90%.In some embodiments, the size resolution of separation is less than approximately 10 μ m.In some embodiments, cell is less than approximately separating in 1 minute.
In some embodiments, system described herein, device, method and product relate to the miniflow platform, its controlled operation that biocompatible ferrofluid is used for particulate and living cells with separate fast.In some embodiments, system as herein described, device, method, and product can carry out by biocompatible concentrated ferrofluid the high flux operation, unmarked classified and separated of cell.The biocompatibility of ferrofluid is based on ionic surface active agent, for example effective balance of citrate, or concentration.Biocompatibility needs neutral pH usually, enough osmotic pressure on cell, and stable ferrofluid (it is unstable that for example, too many ion content can make suspending liquid).This platform utilizes the difference of particle size, shape, elasticity, form etc., to realize fast and effectively to separate.Use the separation based on size of microballoon to show less than for example approximately in 45 seconds, approximately 99% separation efficiency and inferior 10 μ m resolution.System as herein described, device, method and product also provide the erythrocytic continued operation and separating based on shape of living from sickle cell and bacterium.Ferromagnetic microfluid system described herein, device, method is separated and target cell is delivered to sensor array by quick in raji cell assay Raji with product, significantly reduces incubation time and increases diagnostic sensitivity.
Therefore, in some embodiments, disclose and separated the multiple system that is suspended in the target substance in biocompatible ferrofluid.System comprises microchannel, it comprises at least one sample inlet and at least one outlet, microchannel has the length of extending between at least one sample inlet and at least one outlet, microchannel is configured to accept basically continuous sample flow from least one sample inlet, passage is configured to make sample to flow at least one outlet along passage length, and sample comprises plurality of target material and biocompatible ferrofluid.System also comprises a plurality of electrodes, wherein microchannel length is through nearest at least a portion of a plurality of electrodes, and power configuration is for controllably applying at least a electric current to a plurality of electrodes,, with the controlled generation magnetic field model of at least a portion passage length along microchannel, cause at least two kinds of separation in the plurality of target material in sample.
The embodiment of system can comprise any feature described in the disclosure, comprises any following feature.
Power supply can be configured to additionally controllably apply at least a electric current to produce magnetic force component and magnetic moment component.
The plurality of target material that separates can with microchannel in basically separate on the direction of the direction perpendicular of continuous sample flow.
Basically continuous sample flow can for example be passed through, forcing pump, syringe pump, peristaltic pump, vacuum plant, gravity, and/or at least a of capillary force provides.
Power supply for example can be configured to basis, at least a electric current that applies of selected amplitude, selected frequency and/or selected phase place.The separation of at least two kinds of target substances of sample can be at least partly based on one or more of the selected amplitude of for example electric current, selected frequency and/or selected phase place.
A plurality of interelectrode intervals can increase gradually along its length.
Interval between at least some of a plurality of electrodes can be reduced gradually along its length.
A plurality of electrodes can be arranged in electrode layer.
A plurality of electrodes can be arranged in a plurality of electrode layers.
A plurality of electrode layers can perpendicular mode arrangement.
A plurality of electrode layers can concentrically ringed mode arrangement.
The wall of microchannel can limit for example small-sized, carinate, reeded, the fluted or zone that tilts.
Nearest at least a portion that the length of microchannel can be passed a plurality of electrodes with the about angle between the 0-90 degree.
The plurality of target material can comprise at least a material based on cell.Can comprise following one or more based on the material of cell, for example, leucocyte, red blood cell, tumour cell and/or based on the cell of bacterium.
In some embodiments, disclose and separated the multiple device that is suspended in the target substance in biocompatible ferrofluid.Device comprises microchannel, comprise at least one sample inlet and at least one outlet, microchannel has the passage length that extends between at least one sample inlet and at least one outlet, microchannel is configured to accept basically continuous sample flow from least one sample inlet, and is further configured to and makes sample flow at least one outlet along passage length.Sample comprises plurality of target material and biocompatible ferrofluid.Device also comprises a plurality of electrodes of the contiguous microchannel in position, and a plurality of electrodes are configured to when electric current is applied to a plurality of electrode, and generation is along the magnetic field model of at least a portion of the passage length of microchannel.Magnetic field model is configured to cause at least two kinds of separation of plurality of target material in sample flow when sample flow is advanced along at least part of microchannel.
The embodiment of device can comprise any feature described in the disclosure, comprises any above-mentioned feature relevant with system and feature hereinafter described.
Device may further include power supply, is configured to controllably apply electric current controllably to produce magnetic field model to a plurality of electrodes.
A plurality of electrodes can be configured to produce controlled magnetic force component and controlled magnetic moment component.
A plurality of electrodes can be configured to cause at least two kinds of target substances with microchannel in basically separate on the direction of the direction perpendicular of continuous sample flow.
Device may further include and is configured to produce the stream generation unit of continuous stream basically.Stream generation unit can comprise following at least a, for example, and forcing pump, syringe pump, peristaltic pump, vacuum plant, the structure that makes it to flow by gravity, and/or produce the device of capillary force.
Biocompatible ferrofluid can comprise that appropriate ionic species promotes the continuation of cell with the osmotic pressure on the control cell.Biocompatible ferrofluid can comprise the approximately citric acid salt concentration of 5-200 mM.Biocompatible ferrofluid can comprise the approximately citric acid salt concentration of 40 mM.Biocompatible ferrofluid can comprise the ionic strength of the approximately 150mM of design, thereby biocompatible ferrofluid is that grade is oozed and is fit to keep eukaryotic alive.
Biocompatible ferrofluid can have approximately 7.4 pH.
Described at least two kinds of materials can the based target size separate.
Described at least two kinds of materials can the based target shape separate.
Described at least two kinds of materials can the based target resilient separation.
Described at least two kinds of materials can the based target form fractionation.
Described at least two kinds of materials can be hunted down based on following one or more, for example, and the frequency of electrode separation, the electric current that applies and/or the phase place of the electric current that applies.
In some embodiments, the method for separating the plurality of target material is disclosed.The entrance that method is included in microchannel accepts to comprise the continuous sample flow basically of the plurality of target material that is suspended in biocompatible ferrofluid, this sample is passed through along microchannel, and to the electrode of a plurality of positions adjacent channel, apply at least a controlled electric current.Current arrangements is for along at least a portion passage length of microchannel, controllably producing magnetic field model, and is separated to cause at least two kinds of plurality of target material in sample.
The embodiment of method can comprise the described any feature of the disclosure, comprises above-mentioned feature that any and system are relevant with device and feature hereinafter described.
At least two kinds of may further include in the plurality of target material that sorting separates of method enter at least one output channel, and it is based on following at least a difference, for example, and cell size, shape, elasticity, and/or form.
Applying at least a controlled electric current can comprise and controllably apply at least a electric current to produce magnetic force component and magnetic moment component.
To a plurality of electrodes controllably apply electric current cause at least two kinds of the plurality of target material with microchannel in basically separate on the direction of the direction perpendicular of continuous sample flow.
Accepting basically continuous sample flow comprises using and following at least aly from external source, produces pressure so that continuous sample flow to be provided, for example, forcing pump, syringe pump, peristaltic pump, vacuum plant, can produce the structure of the auxiliary pressure of gravity, and/or produce the device of capillary force.
Controllably applying at least a electric current can comprise according to following one or more and apply electric current, for example, selected amplitude, institute's selected frequency and/or selected phase place, wherein the separation of at least two of sample kinds of target substances is one or more of at least part of selected amplitude, institute's selected frequency and/or selected phase place based on for example electric current.
In some embodiments, disclose and concentrated the device that is suspended at least a target substance in biocompatible ferrofluid.Device comprises microchannel, is configured to accept to comprise the sample of at least a target substance and biocompatible ferrofluid, and at least a target substance in the sample of acceptance is concentrated in the inlet flow with relevant input width substantially.Equipment also comprises at least two electrodes, the contiguous microchannel in position, and described at least two electrodes are configured at least when controlled electric current puts on two electrodes, produce controlled magnetic force in comprising the sample of ferrofluid.The controlled magnetic force that produces causes at least a target substance to focus in the stream that produces, and its width is narrower than the relevant input width of inlet flow.
The embodiment of device can comprise the described any feature of the disclosure, comprises any above-mentioned feature relevant with system, first device and method and feature hereinafter described.
At least two electrodes of the contiguous microchannel in described position can be configured to conduct the electric current of controlled supply and produce controlled magnetic force, and this is the physical characteristics of at least two electrodes of basis at least partly.Described at least two electrodes comprise having following at least a structure, for example, the substantially parallel layout of one or more corrugated shape, at least two electrodes basically of one or more straight shapes basically of at least two electrodes, at least two electrodes, and/or at least two electrodes location of taper basically, wherein said at least two electrodes are near each other gradually.
Described at least two electrodes can be configured to cause at least a target substance flow into above and space in microchannel between at least two electrodes.
At least two electrodes can be configured to produce at least two magnetic waves of backward-travelling wave field (reverse traveling field), to cause at least a target substance, are concentrated near center, border between the magnetic wave of at least two generations.
At least two electrodes can comprise the electrode of a plurality of substantially parallel layouts.
Described at least two electrodes can comprise the array of electrode, the relatively adjacent electrode of at least some of array the first electrode is tapered oriented arrangement basically, so that the first electrode is configured to move closer to adjacent electrode, electrod-array is configured to produce magnetic force, with the stream that causes at least a target substance that produces up and adjacent electrode between form.
At least two electrodes can be configured at least when the controllable current with dependent phase is applied to described two electrodes, produce controlled magnetic force in sample.The dependent phase of at least a electric current can be different from the dependent phase of another electric current.
At least a target substance comprises at least two kinds of target substances.A plurality of electrodes can be further configured to cause at least two kinds of in the stream of concentrating that produces plurality of target material separated.
Interval between at least two electrodes can increase gradually.
Interval between at least two electrodes can be reduced gradually.
At least two electrodes can be arranged at least one electrode layer.
At least two electrodes can be arranged in a plurality of electrode layers.
At least a portion that the length of microchannel can be passed at least two contiguous electrodes with the about angle between the 0-90 degree.
Described at least a target substance comprises the target substance based on cell.
In some embodiments, the system that is suspended at least a target substance in biocompatible ferrofluid of concentrating is disclosed.Described system comprises microchannel, is configured to accept to comprise the sample of at least a target substance and biocompatible ferrofluid, and at least a target substance in the sample of acceptance is concentrated in the inlet flow with relevant input width substantially.System also comprises at least two electrodes, the contiguous microchannel in position, and described at least two electrodes are configured at least when controlled electric current puts on two electrodes, produce controlled magnetic force in comprising the sample of ferrofluid.The controlled magnetic force that produces causes at least a target substance to focus in the stream that produces, and its width is narrower than the relevant input width of inlet flow.System comprises that further power supply is controllably to apply controlled electric current at least two electrodes.
The embodiment of system can comprise the described any feature of the disclosure, comprises above-mentioned feature that any and the first system, apparatus and method are relevant and feature hereinafter described.
Power supply can be configured to apply electric current, and it comprises selected amplitude separately, selected relevant frequency separately, and/or selected relevant phase place separately.At least a target substance concentrate can be at least partly based on for example, the selected amplitude separately of the electric current that applies, selected frequency and/or selected phase place.
System further comprises the pressure generation unit, so that inlet flow to be provided, the pressure generation unit comprises following at least a, for example, forcing pump, syringe pump, peristaltic pump, vacuum plant, can produce the structure of the auxiliary pressure of gravity, and/or produce the device of capillary force.
Controlled magnetic force can cause the stream of the generation of at least a target substance be pushed into above and space in microchannel between at least two electrodes.
At least two electrodes of the contiguous microchannel in position can be configured to produce at least two magnetic waves of backward-travelling wave field, cause at least a target substance to be concentrated near center, border between the magnetic wave of at least two generations.
At least two electrodes that are configured to produce at least two magnetic waves of relevant backward-travelling wave field can comprise the electrode of a plurality of substantially parallel layouts.
Described at least two electrodes can comprise the array of electrode, comprise relatively adjacent electrode a plurality of first electrodes of tapered oriented arrangement basically, so that the first electrode moves closer to adjacent electrode, electrod-array is configured to produce magnetic force, with the stream that causes at least a target substance that produces up and adjacent electrode between form.
At least two kinds of electrodes can be configured at least when the controlled electric current with dependent phase is applied to two electrodes, produce controlled magnetic force in comprising the sample of ferrofluid.The dependent phase of at least a electric current can be different from the dependent phase of another electric current.
In some embodiments, the method for concentrating at least a target substance in microchannel is disclosed.Described method comprises to be accepted to comprise the sample that is suspended at least a target substance in biocompatible ferrofluid, and at least a target substance in the sample of acceptance is concentrated in the inlet flow with relevant input width substantially.Method also comprises that at least two electrodes of the contiguous microchannel to position controllably apply at least a electric current, to produce controlled magnetic force in the sample comprising the ferrofluid passage.Magnetic force causes at least a target substance to focus in the stream that produces, and its width is narrower than the relevant input width of inlet flow.
The embodiment of method can comprise any feature described in the disclosure, comprises above-mentioned feature that any and system, device are relevant with first method and feature hereinafter described.
Controllably apply at least a electric current and can comprise following one or more of controllably selecting at least a electric current, for example, relevant amplitude separately, relevant frequency separately and relevant phase place separately.At least a target substance concentrate can be at least partly based on, for example, the selected amplitude separately of at least a electric current that applies, selected frequency and/or selected phase place separately separately.
Method may further include the working pressure generation unit inlet flow is provided, the pressure generation unit comprises following at least a, for example, forcing pump, syringe pump, peristaltic pump, vacuum suction device, can produce the structure of the auxiliary pressure of gravity, and/or produce the device of capillary force.
At least two electrodes are configured to conduct the electric current that controllably applies to produce controlled magnetic force, cause the stream of at least a target substance that produces be pushed into above and space in microchannel between at least two electrodes.
In some embodiments, the device of at least a target substance in the test sample is disclosed.Device comprises microchannel, its be configured to accept to comprise at least a target substance and wherein at least a target substance be suspended in the sample of biocompatible ferrofluid wherein, the detecting device of at least a target substance in working sample, and at least two electrodes of the contiguous microchannel in position.At least two electrodes are configured to produce controlled magnetic force when at least a electric current is controlled while being applied at least two electrodes in comprising the sample of ferrofluid, and the controlled magnetic force that produces causes at least a target substance guiding detecting device.
The embodiment of device can comprise any feature described in the disclosure, comprises any above-mentioned feature relevant with system, apparatus and method and feature hereinafter described.
At least two electrodes can comprise electrod-array, and at least some electrodes are arranged the taper location basically with relative adjacent electrode, thereby at least some electrodes move closer to their adjacent electrode.
At least two electrodes can be configured at least when the controlled electric current that comprises dependent phase is applied to two electrodes, produce controlled magnetic force in comprising the sample of ferrofluid.The dependent phase of at least a electric current can be different from the dependent phase of another electric current.
Sample can comprise the plurality of target material, and device may further include the electrode group, be configured to when controlled electric current is applied to this electrode group, along at least a portion of microchannel length, produce controlled magnetic field model, cause at least two kinds of plurality of target material in sample separated.
Microchannel can be configured to accept the sample flow of from external sample, originating.
Detecting device can comprise the electrode at a pair of interval of measuring the electric capacity in microchannel, and determines the evaluation unit of the existence of at least a target substance based on measured electric capacity.
Be configured to determine that variation that evaluation unit that at least a target substance exists can be configured to the electric capacity in the measured microchannel that causes based on the existence due at least a target substance determines the existence of at least a target substance.
Detecting device may further include capture region, comprise and being configured to and the interactional material of one of plurality of target material, capture region is positioned at the microchannel downstream of the electrode pair at interval, and another electrode to interval is arranged in the downstream of capture region to measure the electric capacity of microchannel.Identify that unit can be configured to determine the initial number of at least a target substance in the electrode pair at interval based on the electric capacity that the electrode pair at interval is measured, determine the end number of at least a target substance with the electric capacity based at another place of electrode to interval, measuring, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
Detecting device can comprise the test set that the order of series connection arranges, the test set of the order setting of each series connection comprises that the electrode at first pair of interval is to measure the electric capacity in microchannel, capture region comprises and being configured to and the interactional material of one of plurality of target material, capture region is positioned at the downstream of the electrode at first pair of interval, and the electrode at second pair of interval is arranged in the downstream of capture region to measure the electric capacity of microchannel.Device can also comprise the evaluation unit, determine the initial number of at least a target substance at the electrode place at first pair of interval with the electric capacity of measuring based on the electrode place at first pair of interval at each test set place, determine the end number of at least a target substance at the electrode place at second pair of interval with the electric capacity that electrode place based at second pair of interval is measured, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
Detecting device can comprise the electrode at a pair of interval of measuring the impedance in microchannel, and determines the evaluation unit of the existence of at least a target substance based on measured impedance.
Be configured to determine that variation that evaluation unit that at least a target substance exists can be configured to the impedance in the measured microchannel that causes based on the existence due at least a target substance determines the existence of at least a target substance.
Detecting device may further include capture region, comprise and being configured to and the interactional material of one of plurality of target material, capture region is positioned at the microchannel downstream of the electrode pair at interval, and another electrode to interval is arranged in the downstream of capture region to measure the impedance of microchannel.Identify that unit can be configured to determine the initial number of at least a target substance in the electrode pair at interval based on the impedance that the electrode pair at interval is measured, determine the end number of at least a target substance at another place of electrode to interval with the impedance based at another place of electrode to interval, measuring, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
Detecting device can comprise the test set that the order of series connection arranges, the test set of the order setting of each series connection comprises that the electrode at first pair of interval is to measure the impedance in microchannel, capture region comprises and being configured to and the interactional material of one of plurality of target material, capture region is positioned at the downstream of the electrode at first pair of interval, and the electrode at second pair of interval is arranged in the downstream of capture region to measure the impedance of microchannel.Device can also comprise the evaluation unit, determine the initial number of at least a target substance at the electrode place at first pair of interval with the impedance of at each test set place, based on the electrode place at first pair of interval, measuring, determine the end number of at least a target substance at the electrode place at second pair of interval with the impedance that electrode place based at second pair of interval is measured, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
In some embodiments, the system of at least a target substance in the test sample is disclosed.system comprises microchannel, be configured to accept to comprise at least a target substance and at least a target substance and be suspended in the sample of biocompatible ferrofluid wherein, determine the detecting device of at least a target substance in sample, at least two electrodes of the contiguous microchannel in position, at least two electrodes are configured at least when controlled at least a electric current puts on two electrodes, produce controlled magnetic force in comprising the sample of ferrofluid, the controlled magnetic force that produces causes at least a target substance to be directed to detecting device, with controllably apply the power supply of controlled at least a electric current at least two electrodes.
The embodiment of system can comprise any feature described in the disclosure, comprises any above-mentioned feature relevant with system, apparatus and method and feature hereinafter described.
The power supply that controllably applies controlled at least a electric current at least two electrodes can be configured to apply the electric current with relevant selected amplitude separately, selected frequency and selected phase place, with at least a target substance guiding detecting device, is wherein at least partly based on the electric current that applies selected amplitude, selected frequency and selected phase place separately.
System further comprises the pressure generation unit, with sampling stream, the pressure generation unit comprise following one or more, for example, forcing pump, syringe pump, peristaltic pump, vacuum suction device, can produce the structure of the auxiliary pressure of gravity, and/or produce the device of capillary force.
In some embodiments, the method for at least a target substance in the test sample is disclosed.Method is included in microchannel accepts to comprise the sample that is suspended at least a target substance in biocompatible ferrofluid, at least two electrodes of contiguous microchannel controllably apply at least a electric current to position, cause at least a target substance to be directed to detecting device to produce controlled magnetic force in the sample comprising the ferrofluid passage, and based on the measurement that detecting device carries out the sample in microchannel, determine at least a target substance in sample.
The embodiment of method can comprise any feature described in the disclosure, comprises any above-mentioned feature relevant with system, apparatus and method and feature hereinafter described.
Controllably apply at least a electric current and can comprise following one or more of controllably selecting at least a electric current, for example, each autocorrelative amplitude, each autocorrelative frequency and/or each autocorrelative phase place.At least partly based on the selected amplitude separately of at least a electric current that for example applies, selected frequency and/or selected phase place with at least a target substance guiding detecting device.
Method may further include the working pressure generation unit and is provided at the sample of accepting in microchannel, the pressure generation unit comprises following at least a, for example, forcing pump, syringe pump, peristaltic pump, vacuum suction device, can produce the structure of the auxiliary pressure of gravity, and/or produce the device of capillary force.
Detecting device can comprise the electrode pair at the interval of measuring the electric capacity in microchannel, and determines that at least a target substance can comprise the existence of determining at least a target substance based on measured electric capacity.
Detecting device may further include capture region, comprise and being configured to and the interactional material of one of plurality of target material, capture region is positioned at the microchannel downstream of the electrode pair at interval, and another electrode to interval is arranged in the downstream of capture region to measure the electric capacity of microchannel.The existence of determining at least a target substance in sample can comprise, determine the initial number of at least a target substance in the electrode pair at interval based on the electric capacity that the electrode pair at interval is measured, determine the end number of at least a target substance at another place of electrode to interval with the electric capacity based at another place of electrode to interval, measuring, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
Detecting device can comprise the test set that the order of series connection arranges, the test set of the order setting of each series connection comprises that the electrode at first pair of interval is to measure the electric capacity in microchannel, capture region comprises and being configured to and the interactional material of one of plurality of target material, capture region is positioned at the downstream of the electrode at first pair of interval, and the electrode at second pair of interval is arranged in the downstream of capture region to measure the electric capacity of microchannel.Determine that at least a target substance can be included in the electric capacity of measuring based on the electrode pair at interval at each test set place and determine the initial number of at least a target substance at the electrode place at first pair of interval, determine the end number of at least a target substance at the electrode place at second pair of interval with the electric capacity based at another place of electrode to interval, measuring, and at each test set place, whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
Detecting device can comprise the electrode pair at the interval of measuring the impedance in microchannel, and determines that at least a target substance can comprise the existence of determining at least a target substance based on measured impedance.
Detecting device may further include capture region, comprise and being configured to and the interactional material of one of plurality of target material, capture region is positioned at the microchannel downstream of the electrode pair at interval, and another electrode to interval is arranged in the downstream of capture region to measure the impedance of microchannel.The existence of determining at least a target substance in sample can comprise, determine the initial number of at least a target substance in the electrode pair at interval based on the impedance that the electrode pair at interval is measured, determine the end number of at least a target substance at another place of electrode to interval with the impedance based at another place of electrode to interval, measuring, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
Detecting device can comprise the test set that the order of series connection arranges, the test set of the order setting of each series connection comprises that the electrode at first pair of interval is to measure the impedance in microchannel, capture region comprises and being configured to and the interactional material of one of plurality of target material, capture region is positioned at the downstream of the electrode at first pair of interval, and the electrode at second pair of interval is arranged in the downstream of capture region to measure the impedance of microchannel.Determine that at least a target substance can be included in the impedance of measuring based on the electrode pair at interval at each test set place and determine the initial number of at least a target substance at the electrode place at first pair of interval, determine the end number of at least a target substance at the electrode place at second pair of interval with the impedance based at another place of electrode to interval, measuring, and at each test set place, whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
Except as otherwise noted, all technology used herein and scientific terminology have such as usually or tradition the identical meaning of understanding.As used herein, article " one of a() " and " one of an() " refer to the grammar object of one or more than one (that is, at least one) article.For example, " element " meaning is an element or more than an element.As used herein, " approximately " when referring to measurable value, during such as amount, duration etc., refer to comprise occurrence ± 20% or ± 10%, ± 5% or ± 0.1% variation, because in the situation that system described herein, apparatus and method, these variations are fit to.It is to be only for convenient and concise and to the point that the disclosure is mentioned scope in the whole text, should not be construed as the fixing restriction to the embodiment of system described herein, apparatus and method.Therefore, the description of scope should be considered to specifically disclose all possible inferior scope in this scope, and individual other numerical value.For example, the description of scope, for example from 1 to 6 should be considered to specifically disclose inferior scope, such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., and the indivedual numbers in this scope, for example 1,2,2.7,3,4,5,5.3,6 and the increment of any whole and part therebetween.
According to the detailed description of following accompanying drawing, the disclosure other and further object, feature, aspect, and advantage will become better understood.
Description of drawings
With reference to following accompanying drawing, these and other aspect will be described in detail now.
Figure 1A comprises microchannel and the electrode schematic diagram with the device that produces magnetic field.
Figure 1B is the COMSOL simulation of the magnetic field (black arrow) of passing ferromagnetic microfluidic device xsect and magnetic flux density magnitude.
Fig. 1 C is the figure along the force and moment of the calculating on the microballoon of microchannel length.
Fig. 1 D is as the magnetic force of the function calculating of frequency and the figure of magnetic moment.
Fig. 2 A is the figure of example embodiment of optional possible microchannel configuration that comprises the electrode spread of 100,150,200 and 300 μ m spacings.
Fig. 2 B is the figure with example passage of a plurality of entrance and exits.
Fig. 3 A is the schematic diagram that comprises the example multidimension electrode configuration of crossed electrode layer.
Fig. 3 B is the figure of the electrode of concentric circles pattern.
Fig. 4 A implements the installation drawing of continuous stream operation by microchannel.
Fig. 4 B-D is the zoomed-in view of the various piece of installing shown in Fig. 4 A.
Fig. 5 realizes the schematic diagram of another example embodiment of the device of discontinuous separation with continuous stream.
Fig. 6 A-C is that illustration uses continuous stream to carry out the figure of the device of three lock out operation.
Fig. 7 carries out the process flow diagram of the example procedure of discontinuous separation to being suspended in the plurality of target material that suspends in ferrofluid.
Fig. 8 A is the figure that describes cobalt ferrite nanoparticles Size Distribution in ferrofluid.
Fig. 8 B describes the AC magnetic susceptibility of ferrofluid and the figure of demagnetization curve.
Fig. 8 C describes the figure of viable count to citric acid salt concentration.
Fig. 9 A-B comprises the figure that is presented at ferrofluid endoparticle behavior in the situation that has magnetic field.
Figure 10 A-D comprises curve map and the diagram that is presented at ferrofluid endoparticle behavior in the situation that has magnetic field.
Figure 11 A and 11B are the figure that describes the cell separation of all types of target material.
Figure 12 A-C comprises curve map and the diagram that is presented at ferrofluid endoparticle behavior in the situation that has magnetic field.
Figure 13 is the schematic diagram of microfluidic devices, and it comprises the electrode group, and it is configured to produce magnetic field the target substance of sample is focused on basic single file stream.
Figure 14 is configured to target substance to be focused on the alternately schematic diagram of the part of another example microfluidic devices in gap.
Figure 15 is the schematic diagram of part of another example embodiment of microfluidic devices.
Figure 16 concentrates the process flow diagram of the example procedure of target substance in microchannel.
Figure 17 is the schematic diagram of part of another example embodiment of microfluidic devices.
Figure 18 comprises the schematic diagram of capacitance detector and describes the figure of bacterium by near cause the detecting device face capacitance variations.
Figure 19 is the process flow diagram of the example procedure of at least a target substance in test sample.
Figure 20 is the schematic diagram of general-purpose computing system.
In each figure, similar quotation mark represents similar element.
Describe in detail
Herein disclosed is system, method, device, product, with various embodiments, comprise that device (for example, for separating of the plurality of target material that is suspended in biocompatible ferrofluid), it comprises microchannel, this microchannel comprises at least one sample inlet and at least one outlet, and microchannel has the passage length that extends between at least one sample inlet and at least one outlet.Microchannel is configured to accept basically continuous sample flow from least one sample inlet, and is configured to make sample to flow at least one outlet along passage length.Sample comprises plurality of target material and biocompatible ferrofluid.Device also comprises a plurality of electrodes of the contiguous microchannel in position, a plurality of electrodes are configured to when electric current is applied to a plurality of electrode, the magnetic field model that produces along at least a portion of microchannel length, magnetic field model is configured to when sample flow during along the advancing at least partly of microchannel, cause at least two kinds of plurality of target material in sample flow separated.
Also disclose and concentrated the device that is suspended at least a target substance in biocompatible ferrofluid.Device comprises microchannel, be configured to accept to comprise the sample of at least a target substance and biocompatible ferrofluid, at least a target substance in sample of accepting is concentrated in the inlet flow with correlated inputs width substantially, at least two electrodes with the contiguous microchannel in position, at least two electrodes are configured at least when controlled electric current puts on two electrodes, produce controlled magnetic force in comprising the sample of ferrofluid.The controlled magnetic force that produces causes at least a target substance to focus in the stream that produces, and its width is narrower than the relevant input width of inlet flow.
System, device, method are further disclosed, and embodiment, the device that comprises at least a target substance in test sample, device comprises microchannel, its be configured to accept to comprise at least a target substance and wherein at least a target substance be suspended in the sample of biocompatible ferrofluid wherein, the detecting device of at least a target substance in working sample, and at least two electrodes of the contiguous microchannel in position.At least two electrodes are configured at least when controlled at least a electric current is applied to two electrodes, produce controlled magnetic force in comprising the sample of ferrofluid, and the controlled magnetic force that produces causes at least a target substance guiding detecting device.
In general, system described herein, apparatus and method ferrofluid used are the colloid admixtures of the magnetic-particle of nano-scale, vectolite for example, and the covering surfaces activating agent, be suspended in the mounting medium compatible with surfactant material.For example, the example reaction of generation magnetic-particle is as follows:
2?FeCl3?+?FeCl2?+?8?NH3?+?4H2O?Fe3O4?+?8?NH4Cl。
The suspending liquid of the magnetic iron ore of 10 volume % has the approximately saturation magnetization of 560G.The magnetization of each single domain particle is 10 μ s magnitudes to the responsive time constant of highfield.High magnetic field gradients can be used for the detent rail magnetic fluid.Under the condition of this highfield existence, " spike " and other significant features may appear at the ferrofluid surface.
In some embodiments, the particle diameter scope can be from about 1nm to about 100nm, and any whole or increment part therebetween.For example, and without any restriction, the particle diameter scope can be 1-10nm, 1-20nm, 5-50nm or 10-100nm.In some embodiments, the about 10nm of particle diameter average out to.Volume fraction can from 0.1% to approximately 10%, and any whole or increment part therebetween.
In some embodiments, ferrofluid described herein is biocompatible, and living cells was kept several hours, and physical property or continuation are not degenerated, and can extend the detection of target sample.Biocompatible ferrofluid goes for keeping cell type and/or the shape of any work, for example any animal or plant histocyte type, any microorganism, or its any combination.Ferrofluid can also be suitable for the suspending particle of any type, and the particle of any size or shape, or cluster of particle or clump, no matter live or non-work.
Citrate is effective surfactant in ferrofluid, and its major part in cell culture is also compatible.Therefore, in some embodiments, citrate is used for the stabilization ferrofluid and provides making cell survival ionic medium wherein.Under this background, determine newly with citric acid salt concentration optimum, to wish because very little or too many citrate can cause particle aggregation and precipitation in ferrofluid.And the survival of ferrofluid inner cell depends on that having enough ionic species promotes the continuation of cell with the osmotic pressure on the control cell.In an example embodiment, causing in the ferrofluid of the stable soliquid of magnetic nanoparticle citric acid salt concentration is about 40mM.And than the homocitric acid salinity, may start to reduce gradually ferrofluid stability, any position that the concentration of citrate can be between 5-200mM, and any whole or part increment therebetween, depend on feature and the type of ferrofluid used.(describe in Fig. 8 C) in example embodiment, also determined with the stable pH approximately 7.4 that obtains of citric acid, the least concentration that at first causes the citrate of most cells survival during several hours is 40mM approximately.Under this ion concentration, cell has vigor and ferrofluid to stablize.Therefore, in some embodiments, approximately the citric acid salt concentration of 40mM can be used as effective biocompatible ferrofluid.As this paper expectedly, and depend on the type of the ferrofluid that uses, ferrofluid described herein can also be stabilized in following pH scope: about 2-11, and any whole or part increment therebetween.In some embodiments, ferrofluid is biocompatible, thereby cell can be survived at least about 1 hour, approximately 2 hours, approximately 3 hours, approximately 5 hours, approximately 10 hours, high to approximately 24 hours or even longer in ferrofluid.In some embodiments, biocompatible ferrofluid used comprises the biocompatible ferrofluid of design, have the approximately ionic strength of 150mM, and this biocompatible ferrofluid be configured to etc. to ooze and be suitable for keeping eukaryotic alive such as people's cell, people's cell etc.
1. piece-rate system and device
As described herein, the unmarked operation and the micro-fluidic system that separates that are used for bio-compatible ferrofluid target substance (for example, cell and microorganism) based on the flowing molten iron body dynamics are provided.In some embodiments, system comprises the ferrofluid based on water as the interior even magnetic environment around cell or other particle of microchannel.Ferrofluid inner cell and other non-magnetic particle be similar to the mode of electron aperture in semiconductor serve as " magnetic space " (Kashevsky, 1997,
Phys Fluids9:1811-1818).The magnetic field gradient that outside applies can attract magnetic nanoparticle, and this causes non-magnetic particles or cell effectively to be pushed open (Rosensweig RE (1997)
Ferrohydrodynamics(Dover, New York); Odenbach S (2002)
Ferrofluids:Magnetically Controllable Fluids and TheirApplications(Springer, New York)).Recently, this principle be applied to catching the non magnetic microballon between the thin magnetic film island in the microchannel of filling ferrofluid (Yellen et al., 2005,
Prot Natl Acad Sci USA102:8860-8864).In contrast, system of the present disclosure, apparatus and method comprise the microfluidic devices with electrode (can be randomly Integrated electrode, for example integrated copper electrode), and its transmission current produces and can design/configurable magnetic field gradient with part.Following appendix A and B provide owing to microchannel (sample that comprises target substance to be processed flows by it), applying magnetic field and have caused mathematical description and the analysis of the behavior that occurs and interactive relation.
The example embodiment of Figure 1A-1D explaination iron microfluidic devices and particle operating platform.Figure 1A comprises that microchannel 110 and following electrode 120(draw not in scale) the schematic diagram of device 100.In some embodiments, in some embodiments, the sinusoidal current that provides 90 ° of relative to each other phase place lockings from two output channels of amplifier (for example,
I 1 With
I 2 ).Adjacent electrode on base material arranges in a certain way/connects, with the orthogonal manner transport current and support travelling-magnetic-field in microchannel.Non magnetic microballoon or cell that the magnetic field gradient that produces promotes in the iron microchannel rise and enter interelectrode gap.Traveling-wave field also causes rotary-top and the rolling of cell along passage, causes along this passage length with the continuous translation of the frequency higher than threshold value.The particle movement that causes can be observed from top with upright microscope 130, and, for example use the CCD camera with per second 18 frame-grabs, be used for further analyzing.Iron microfluidic devices and further describing of particle operating platform are provided at, for example, on Dec 7th, 2010 submitted to, PCT application number PCT/US10/59270, exercise question is the U.S. Provisional Application sequence number 61/267 of " n cell that is undertaken by biocompatible ferrofluid operates and sorting " and submission on Dec 7th, 2009, in the U.S. Provisional Application sequence number 61/407,738 of submitting on October 28th, 163 and 2010, its all the elements this by reference integral body incorporate into.
Continuation is with reference to Figure 1A, and electrode 120 can be arranged in electrode layer, and the contiguous microchannel 110 in position, can be positioned at above base material, for example the insulated metal base material 150 of standard.For example, can use the aluminium base of coated insulating polymer, it can dispel the heat efficiently, and for example AC electric current of 10A is conducted by electrode under low-voltage.In an example embodiment, can use the single electrode layer, describe as Figure 1A.In some embodiments, the multi-electrode layer is used for providing multidimensional to control (thereby with the multidimensional that can realize magnetic field, control, described magnetic field can be produced and put on sample in microchannel with the target substance in operation and sample separation).For example, Fig. 3 A has described the example of the multidimension electrode configuration that comprises the crossed electrode layer.
In given electrode layer, electrode can be that approximately 30 μ m are high, and approximately 300 μ m are wide and approximately 2cm is long.In optional embodiment, give the electrode in given layer can be approximately 5-100 μ m is high, approximately 0.01-1mm is wide, and the about long any range of 0.1-10cm, and any whole or part increment therebetween.Therefore, will be appreciated that, the size of electrode used therein is restriction not, and can change between a plurality of electrode layers.And electrode can comprise any shape, curvature or pattern, and can comprise the variable gap size between electrode.For example, Fig. 2 A has described the electrode spread at 100,150,200 and 300 μ m intervals in passage area.As described, in some embodiments, can use the multilayer orthogonal modes, for example, as described in Fig. 3 A.In an example embodiment, can use concentric electrode pattern as described in Fig. 3 B so that the row wave energy more effectively moving particles or cell enter round relevant range, leave round relevant range or the different piece IT of circle relevant range they.In some embodiments, electrode can have " wavy " (bending), or is generally crooked shape, in order near particle or cell are introduced the interference force and moment.These wavy zones in fact can be for even, inhomogeneous, random and/or periodic, and can be dispersed throughout electrode.Should be noted that, in a given electrode layer, the shape of electrode, interval and pattern may change, and can further at a plurality of interlayers, change, make any combination of shape, size, interval and pattern to occur at electrode layer with across multilayer, (these attributes are further controlled, or configurable to have the magnetic field of the hope of desirable attribute with generation, for example, be applied to electric current on electrode by control).
In some embodiments, electrode can comprise any suitable conductive material, for example copper.In some embodiments, electrode can be made by the copper layer of the printed circuit board of photoresist mask wet etching thermal envelope (on the insulated metal base material).Be noted that any etching type or other suitable method for making can be for generation of electrodes.
Passage (for example microchannel shown in Figure 1A 110) can comprise at least one entrance and at least one outlet, and can be configured to make the length of passage at least a portion by the vicinity of electrode, and at least a portion of can thereby cross electrode.For example, do in some embodiments, microchannel can rotate approximately 90 degree, thereby the electrode of device is basically parallel with its length.In some other embodiments, passage, with the angle of approximately between the 0-90 degree and therebetween any whole and increment part, crosses electrode.In further example embodiment, passage crosses electrode with basic straight line.In other other example embodiment, passage crosses electrode with arc, bending or common irregular pattern.
In some embodiments, the microchannel scope can be for 20-100 μ m is high, 1-3 mm is wide and 2-3 cm is long, and any whole or part increment therebetween.Can use other size value for passage.In some embodiments, passage can comprise pocket, ridge, trough and/or the inclined-plane of the interior any number of conduit wall and size, thereby the particle of advancing in passage or cell can be based on conformational effect concentration of local or the dispersions of conduit wall profile.Additional examples with passage of a plurality of entrance and exits is described in Fig. 2 B.Passage can comprise any suitable material.For example, in some embodiments, passage can pass through the soft lithography stamp, from dimethyl silicone polymer (PDMS) preparation, and with the thin PDMS insulation course of coated electrode, be connected (Mao et al., 2006,
Nanotechnology17:3447).In some embodiments, channel height can be selected lower than local flowing molten iron body dynamics stream optimal value, in order to minimize its potential effect for particle moving.
Although do not need, before ferrofluid/mixture of microspheres was imported microfluidic devices, passage can clean approximately 10 minutes with 1% triton-X solution, with the particle that minimizes the PDMS wall, adhered to.Will be appreciated that, replacedly, base material, insulation course and passage can comprise the material with similar characteristics and/or character separately.Therefore, in order to define electrode, device described herein and system can be built upon on the cheap printed circuit board that for example is characterized as by the etched insulated copper layer of the transparent mask of single low resolution.As described, microchannel can be built by the soft lithography that uses the low resolution mould.In some embodiments, device is made does not need clean room, thereby and, can make fast and at an easy rate.
In order to operate sample so that sample (is for example carried out desirable processing, separate the plurality of target material at least two kinds, concentrate target substance etc.), by apply to electrode (electrode 120 of for example describing in Figure 1A) controlled at least a electric current from power supply (and, in some embodiments, at least two kinds of electric currents) produce the magnetic field of advancing in passage, to create along the magnetic field model of at least a portion of microchannel length.Can be applied to how about 7A peak-peak amplitude and from the about alternating current of the frequency of 10Hz to 100kHz to electrode, the about maximum magnetic field strength of 90 Oe in this corresponding ferrofluid.In some changed, the magnetic field intensity scope that produces can be 1-200 Oe and any whole or increment part therebetween.In an example, magnetic field is to produce by the alternating current that applies quadrature to single-layer electrodes, to create the periodic magnetic field mode of along microchannel length, advancing.According to this configuration, device can be created the magnetic field gradient that causes the time average power of cell or particle, and the magnetized local rotation of ferrofluid, and it finally causes the moment on non-magnetic particle, as shown in Figure 1B.Figure 1B describes magnetic field (black arrow) that moment preset time passes ferromagnetic microfluidic device xsect and the COMSOL simulation of magnetic flux density magnitude.More shallow arrow is described the magnetic field of interior every 30 ° of one-period.Shown analog result is being carried out under the input of 12-A peak-to-peak current under 1670 Hz frequencies.
When controlled at least a electric current was applied to electrode, two or more target substances (cell or particle),, at least partly due to magnetic force, pushed to channel roof from electrode, and there at least partly due to magnetic moment, they start along its length rotation and rolling.The magnetic susceptibility of the frequency dependence of the specific iron magnetic fluid that uses can thereby be simulated in the device behavior.For given particle size, its speed can depend on the local force and moment value along passage length, for example, (be presented under 4.6kHz under the peak-peak input excitation of 7-A, on the microballoon of 6-μ m diameter along the figure of the force and moment of the calculating of microchannel length) as shown in Figure 1 C.
Under low frequency, magnetic force component is usually leading, promotes non-magnetic particles upward to channel roof (that is, the surperficial relative surface of adjacent electrode), and enters interelectrode space.Under high-frequency, the particulate of rolling can overcome the repulsion that is reducing that magnetic force causes, and moves along passage continuously, as shown in Fig. 1 D.Fig. 1 D is as the magnetic force of frequency function calculating and the figure of magnetic moment (the input current amplitude that is used for simulating is the 7A peak-peak, supposes that the sliding rate for all simulations is 1) for the same particle between electrode on channel roof.
For example, the typical magnetic force that can be applied on the particle of several micron diameters can be in tens skin newton's magnitude, and it is significantly greater than light tweezer typical magnetic force on the micron-scale particle.In some embodiments, driving force can increase by applying larger exciting current.For example, the simple heat radiator content peak-peak input current of 10-A (Mao L, Koser H (2006) the Toward ferrofluidics for p-TAS and lab on-a-chip applications. at the most that can at room temperature keep passage
Nanotechnotogy17:34-47).
Therefore, attribute (intensity, frequency, phase place etc.) by applying from the magnetic field that at least a electric current produces to electrode (such as electrode 120) can based on, the special character (for example, amplitude, frequency, the phase place of the electric current that applies) of the electric current that for example, applies by electrode.In some embodiments, the attribute that is applied at least a electric current of electrode can use controller to control, for example based on the controller of processor, the controller 160(that for example describes in Figure 1A or some other calculation elements), it can be defined as producing and keeping the current properties of the required needs in some required magnetic fields.For example, in some embodiments, controller can make current properties (for example carry out the dynamics adjustment based on the condition that detects in device, if determine the non magnetic target substance in sample, particle for example, cells etc. do not have as required or expection is processed, and for example they are not suitably separated, and change magnetic field).
Extraly, and/or alternatively, in some embodiments, the attribute in the magnetic field that produces can also be based on the configuration of microchannel (for example, be used for implementing its structure, the material of passage), the configuration of electrode (for example, the spatial relationship of their layout, the relative microchannel of electrode, be used for to implement the material of electrode etc.), and other factors.Therefore, in order to control the magnetic field that produces, for example install 100 separation function with control and adjusting, one or more elements of component devices can be controlled or be operated.For example, as described, in some embodiments, be applied to sample so that in sample in the plurality of target material at least two kinds of separated magnetic fields of energy can by to electrode (for example electrode 120 of Figure 1A), controllably applying at least a electric current, (by power supply, be provided, its can with tripping device, for example install 100 and connect) control.In some embodiments, controllably apply at least a electric current to device 100 electrode and can comprise and control/arrange power supply, thereby power supply applies and has at least a electric current of selected amplitude, selected frequency and/or selected phase place separately.Under those environment, the separation of at least two kinds of target substances of sample can be at least partly based on, one or more of the selected amplitude separately of the electric current that applies, selected frequency and selected phase place.
As described, in some embodiments, produce and put on the attribute in the magnetic field of sample and can be for example according to the configuration of the electrode of the contiguous microchannel of configuration and/or position of microchannel, determine.For example,, referring again to Fig. 2 A, shown the schematic diagram of example embodiment of the optional possible microchannel configuration of the electrode spread that comprises 100,150,200 and 300 μ m spacings in passage.Fig. 2 B has described approximately between 0.17-0.19cm, and the passage with 4 entrances and 4 outlets is arranged.In another example, and referring again to Fig. 3 A and 3B, shown the schematic diagram of describing the optional embodiment of electrode mode.Particularly, Fig. 3 A has described the multi-layered electrode pattern of quadrature basically, and Fig. 3 B describes the electrode of concentric circles pattern.Thereby each configuration of describing in Fig. 2 and 3 provides another kind of method/mechanism control magnetic field properties, and it can be produced for example to separate, concentrate, guide and/or operate in addition and drive the target substance (for example passage 110 of Figure 1A) in sample mobile in microchannel.
In some embodiments, tripping device, the device 100 of Figure 1A for example, can be configured to separate with continuous stream and be delivered to target substance in the sample of microchannel of device.Referring now to Fig. 4 A, Fig. 4 A has shown the figure of the continuous stream tripping device 200 of display part, uses and is configured to make the microchannel that in sample, target substance flows to realize, the magnetic field that produces by electrode is applied thereto.The device 100 that device 200 is described in can similar Figure 1A, perhaps it can be based on having, for example the different embodiments of different electrode configurations and/or the configuration of different microchannel.Fig. 4 A has described and has allowed the sample 240 access to plant entrances that suspend in ferrofluid 250 and by separation chamber, and the continuous stream device that leaves through a plurality of outlets of the particle that is configured to catch special size.Particularly, device 200 comprises entrance (also referred to as the entrance stage) 210, separation chamber 220 and outlet (also referred to as the outlet stage) 230.In the example of Fig. 4 A, comprise two kinds of different target materials, namely the continuous stream sample of 2 μ m particles and 5 μ m particles, import by entrance 210.For example, in some embodiments, sample can the working pressure pump, at least a importing of the device of syringe pump, peristaltic pump, vacuum plant and/or any other coupling entrance 210, causes the entrance of sample 240 accesss to plant 200 and flows to separation chamber 220 and outlet 230.Extraly and/or alternatively, in some embodiments, continuous stream can be passed through gravity (for example, the difference in height between the entrance and exit pond) or by capillary force, realize.Show the zoomed-in view of the sample that comprises two kinds of target substances in Fig. 4 B.
Apply controlled electric current by installing 200 special electrode configuration, the magnetic field with association attributes (at least partly based on the controlled electric current that applies by electrode) produces, and causes target particles to separate.Normally, the sample (comprising target substance) that is separated in of particle carries out while flowing through separation chamber 220.Zoomed-in view as Fig. 4 A and Fig. 4 C(demonstration separation chamber) as shown in, in the example of describing in Fig. 4 A and 4C, the magnetic field that produces causes 2 μ m particles and 5 μ m particle separation, and the latter is hunted down in center clearance stream.Subsequently, and as shown in Fig. 4 A and 4D, 2 μ m flow to the outlet A in outlet stage 230, and 5 μ m grain flows are to the outlet B in outlet stage 230.Remaining sample flow is to waste outlet C.Thereby the device 200 of describing in Fig. 4 A-D is suitable for separation and the sorting of two or more grain types, based on a kind of of size, shape, elasticity, form etc. or combination.
, with reference to Fig. 5, shown the schematic diagram of another example embodiment of realizing the device 300 of discontinuous separation with continuous stream.Device 300 comprises one or more entrances, for example, accepts the entrance 310 of ferrofluid and cell sample, and accepts two entrances 312 and 314 of ferrofluid.In some embodiments, can use the entrance of any number, each can be configured to accept will be by the ferrofluid of device processing and/or the potpourri of target substance, and for example the controlled of magnetic field by the guiding of the microchannel at device applies.In the embodiment of Fig. 5, the continuous stream that provides by one or more pressure mechanisms can be provided the material of accepting by entrance, and for example pressure mechanism 302, cause this continuous stream.Explain as this paper, provide the mechanism of continuous stream can comprise following one or more, for example, forcing pump, syringe pump, peristaltic pump, vacuum plant, based on the equipment of gravity, and/or capillary force.In some embodiments, the entrance stage can comprise centralized mechanism, and target substance is focused in single stream, then it process in magnetic field of the microchannel by being applied to device 300.Centralized mechanism can based on fluid mechanics concentrate and/or can according to based on the concentrating of magnetic (for example, with permanent magnet and/or another group electrode be created in entrance be applied near the stage sample).
As Fig. 5 further as shown in, device 300 also comprises microchannel 320, it can be similar to the passage 110 of Figure 1A, thereby and can be various shapes and configuration, and can build a kind of in multiple different materials.Microchannel 320 a plurality of electrode 330(of process are copper electrode for example, or the electrode of other type) contiguous at least a portion.A plurality of electrodes 330 can be configured to have size and other physical attribute,, to promote to produce controlled magnetic field with controlled attribute (when controlled electric current during through electrode), are used for the samples that flow at microchannel 320 are processed.For example, in the embodiment of Fig. 5, the two or more separation that are suspended in the target substance in ferrofluid can be carried out in the magnetic field that produces, and separate the result of a plurality of target substances as magnetic field, and the material of separation is directed at least one outlet pond.As shown in Figure 5, a plurality of electrodes comprise 8 " wavy " electrodes, and its layout (for example, with the various angles of relative microchannel 320) makes when controlled electric current is applied to a plurality of electrode 330, and the magnetic field with direction shown in arrow 350 is produced.Other attribute in the magnetic field that produces can be controlled by at least a electric current that control is applied to a plurality of electrodes.In some embodiments, controlling the electric current that applies can carry out with the controller 160 of controller such as Figure 1A.And by using the difference configuration of electrode, the attribute in magnetic field can further be adjusted/control, and for example, has the magnetic field of different directions with generation.The non magnetic target substance that the magnetic field that operative installations 300 produces causes being suspended in ferrofluid is directed to the zone that basically is limited between electrode 332 and 334.In some embodiments, can use permanent magnet to supplement or to substitute a plurality of electrodes, be applied to the magnetic field on the sample that flows in microchannel with generation.For example permanent magnet can be used for applying and flowing vertical magnetic field gradient,, to promote target substance, promotes the separation of target substance.
The magnetic field of advancing that produces (for example,, by applying of at least a electric current, for example having the applying of two kinds of electric currents of out of phase) causes producing magnetic force and torque, and it is applied to (for example cell, bacterium, other particle) on the plurality of target material.This causes a plurality of target substances to be pushed simultaneously (for example, using magnetic force upward to top with to the interelectrode gap upper space) and uses magnetic moment to cross the width (for example, the nearest surperficial relative surface of ionization electrode) of passage along the top rolling.In high frequency and small electrode gap, magnetic moment leading (cause cell advance the track of traverse gap), in low frequency and/or large gap, magnetic force leading (causing cell to stop concentrates in each track).Thereby the magnetic field that applies can be used for the discontinuous separation of realize target material, because in the device of the clearance gap with increase, cell will finally be limited in separately in track.Magnetic field causes the separation of target substance, and this is based on following one or more, for example, and their size separately, their shape separately, their each own elasticity, their form separately and other characteristic of target substance.
When the sample of magnetic fields is advanced and near outlet during the stage along microchannel length, many kinds of substance separated (for example, according to their size, shape, form, elasticity etc.), and be directed to separately separation outlet pond.For example, in the embodiment that Fig. 5 describes, little target substance (for example, cellule or bacterium) drift about to the left side of device 300, for example, the direction in outlet pond 344, larger target substance (such as leucocyte, red blood cell etc.) drifts about to different directions, for example to outlet pond 342.Other material of sample (for example, waste material) can exit pond 340.In some embodiments, can only use an outlet.
The device 300 of sending continuous sample stream is realized particulate and cell continuous high-throughoutly separates, operation and sorting.In some embodiments, target substance process/operation occur with outside apply flow vertical.In the case, the transhipment of plurality of target material can be not coupling with the mechanism of separating.Flow velocity can be optimized for high flux, and cell separation area size can be correspondingly selected subsequently.The sheath of shearing from side by fluid dynamics flows or passes through magnetic force, and target substance (for example, cell) potpourri can focus on the single gap on adjacent electrode and between adjacent electrode.To continue to separate and sorting cells by the magnetic field of advancing that electric current in electrode produces, when they drag downstream with stream, cross the width of separation chamber.
In some embodiments, by controlling excitation frequency, can realize controlling, wherein target substance will be trapped in initial electrode gap, and target substance is with separated.Gradually change by the gap width between adjacent electrode, also can realize other control of separation parameter.Larger gap makes it more easily catch and concentrate less particulate/cell.For example, the gradient of adjacent segment size (for example, 100 μ m, 200 μ m and 300 μ m) can be separated and then be concentrated respectively 10 μ m, 5 μ m and 2 μ m microballoons, for example, as shown in Fig. 6 A-C, its display device is as installing 300 operation, and the use continuous stream is carried out three separation.In some embodiments, similarly method can be used for creating the independent stream of leucocyte, red blood cell and blood platelet from blood.This will make it may obtain fast and exactly the blood count results within a few minutes.Other target cell (for example tumour cell of circulation) can separate from remaining haemocyte, and this is by making their total fluid dynamics volumes basically be different from haemocyte to separate reliably with microballoon (magnetic or nonmagnetic) mark them.Be different from common separation equipment, flow cytometer for example, typically at the cost of 100,000 dollars of levels, occupy whole experiment table top, and need many assemblies, and comprising a plurality of laser instruments, pump, pipeline, reagent, fluorescent dye and those skilled in the art, discrete microchannel tripping device as herein described and system in minutes (rather than several hours) are completed sample preparation, and in enough cheap packing, it can be processed after processing finishes.
, with reference to Fig. 7, shown the process flow diagram that carries out the example procedure 400 of discontinuous separation to being suspended in plurality of target material in ferrofluid.Program 400 is included in microchannel such as Figure 1A, 4 and 5 in the entrance of the microchannel of describing to be accepted 410 and comprises the multiple sample flow that is suspended in the target substance in biocompatible ferrofluid basically continuously.Biocompatible ferrofluid can be any ferrofluid described herein, is configured to keep biological specimen (for example, human or animal's cell) and reaches the time period (for example a few minutes were by several hours) of relative prolongation.In order to obtain to comprise the continuous sample flow of a plurality of target fluids, use following at least a, for example, forcing pump, syringe pump, peristaltic pump, vacuum plant, can produce the structure of the auxiliary pressure of gravity/device, and/or the device of generation capillary force, the pressure for providing continuous sample to flow from external source is provided.Also can use the device of other type.
At the entrance of the device based on microchannel, accept sample, sample passes through 420 along microchannel.When sample flow during through the microchannel of device, at least a controlled electric current apply 430 electrodes in a plurality of positions adjacent channel (as, electrode, the electrode 330 of for example describing in Fig. 5).Controlled at least a current arrangements is for along at least a portion passage length of microchannel, controllably producing magnetic field model, and is separated to cause at least two kinds of plurality of target material in sample.Especially, as described herein, controlled at least a electric current (and configuration of the configuration of selected electrode and/or microchannel) can be controlled the attribute in generation magnetic field and made magnetic field is programmable or configurable.The magnetic field that produces generally includes magnetic force and magnetic moment component, causes nonmagnetic substance (comprising target substance) to be pushed in ferromagnetic microchannel and rises and enter interelectrode gap.Advancing also causes rotary-top and the rolling of cell along passage, causes along this passage length with the continuous translation of the frequency higher than threshold value.The magnetic field that produces causes different target substances to have different spaces behavior and motion in microchannel, and it depends on size, shape, form, elasticity and the further feature of target substance at least partly.Thereby all types of target material is separated, sorting or be distinguished from each other in addition, makes the different target substance in the sample that is present in continuous flow identified.
Therefore, as described herein, can realize that it merges biocompatible ferrofluid based on the pilot system of stream.In some embodiments, higher flux can separate to realize by carrying out the Biology-iron magnetic fluid, and the stream that flows simultaneously is continuously with the entrance of fresh target substance (for example, cell) introduction channel.In outlet, the pearl that enters, cell, particle are sorted in different exit passageways.Therefrom, cell can collect for detection of or directed outwards or inside (namely integrating) sensor.For example, the stream device allows the sample access to plant entrance that suspends in ferrofluid and by separation chamber, and through a plurality of outlets of the particle that is fit to catch special size, leaves.For example, as described in Fig. 4 A, 2 μ m particles can flow into outlet A, and 5 μ m particles can flow into outlet B etc., and remaining sample can flow into outlet C.In some embodiments, do not need stream to guide cell.On the contrary, excitation can be used for guiding them.And for example, sensor can be integrated directly into along in the pocket on the side of circulation road.
Operation is not only depended on and is also depended on cell shape and elasticity by cell size.For example, bacterium allows them to separate from healthy blood cell with drepanocytic different size, shape and elasticity.In some embodiments, particle separation can depend on size and frequency.In some embodiments, critical frequencies can also depend on electrode gap.For example, at first larger microballoon can be hunted down in less gap.By utilizing this phenomenon, sorting can be carried out based on particle or target size.And system can replace between the operation excitation of institute's selected frequency and another frequency, helps to interrupt possible nanoparticle chains, and it can be because excitation forms.In some changed, " wavy " electrode can be used for preventing that pearl or cell are agglomerated into bulk when flows down channel." wavy " electrode is introduced and is disturbed force and moment to smash reunion on pearl or cell, allow larger indivedual pearls or cell to arrange as the pearl on necklace.Interrupt the chain of nano particle by periodicity, the physical attribute of ferrofluid remains unchanged in time.And target cell can be effectively, rapidly, and in unmarked mode, be concentrated, catch, locate or orientation sensor surface simply.Can be based on size, shape, elasticity, form etc. with cell or grain type exit such as separation method, or, by increasing or reduce interelectrode spacing, based on size, shape, form, elasticity and/or some other characteristic, catch cell or grain type with cell or grain type exit.
The program of realization separating at least one target cell (cell type) from sample is included in suspension cell in the bio-compatible ferrofluid and forms sample, (these electrodes angle of microchannel 0-360o are relatively arranged by crossing a plurality of electrodes to make sample, while for example with the angle of the longitudinal axis 90o of relative microchannel, arranging) microchannel, a plurality of electrodes are basically parallel with the length of microchannel.Program described herein may further include to a plurality of electrodes and applies electric current to create the magnetic field model along microchannel length, and based on cell size, shape and flexible at least a difference sorting cells, enters at least one output outlet.Separation can be passed through effectively, rapidly, and in unmarked mode, concentrates, catches, locates or orientation sensor surface generation simply.
For example, in some embodiments, program described herein can also be based on size isolation of target substances (for example cell and/or particle).This separation based on size can prove to have, for example, approximately 50% efficiency, approximately 60% efficiency, approximately 70% efficiency, approximately 80% efficiency, approximately 90% efficiency, approximately 92% efficiency, approximately 94% efficiency, approximately 96% efficiency, approximately 97% efficiency, approximately 98% efficiency and about 99% separation efficiency.Size resolution in separating treatment can be, for example, be less than approximately 10 μ m, be less than approximately 9 μ m, be less than approximately 8 μ m, be less than approximately 7 μ m, be less than approximately 6 μ m, be less than approximately 5 μ m, be less than approximately 4 μ m, be less than approximately 3 μ m, be less than approximately 2 μ m, be less than approximately 1 μ m, be less than approximately 0.5 μ m, be less than approximately 0.1 μ m and be less than approximately 10 nm.This separation can be being less than approximately 2 minutes, be less than approximately 1 minute, be less than approximately 45 seconds, be less than approximately 30 seconds, be less than approximately 20 seconds and be less than approximately and complete in 10 seconds.
As further described herein, the continued operation of target substance with based on also realizing the separating of shape (for example, cell, the red blood cell of for example living separates with sickle cell and/or bacterium).The ferromagnetic microfluid of this outstanding proof by quick separation with target cell is delivered to sensor array, significantly reduces incubation time and increases the ability of diagnostic sensitivity in raji cell assay Raji.
Micro-fluidic system described herein and device have advantages of that some are unique, because they provide the laminar flow platform to use under the tiny sampler size.Systems/devices further provides rapid diffusion and fast results, can be portable, and can integrate other existing sensor (as describing in further detail hereinafter).For example, system described herein, apparatus and method can be used for the sterilizing of the adult stem that obtains from blood sample, are used for the background of fight soldier and seaman's wound healing and neomorph.System described herein, apparatus and method can also be used for low-level bacterial contamination in blood that fast detecting (for example,＜1 minute) contributes.Under the wound emergency of battlefield, this can be useful especially.System described herein, device, method needing can also to be used for detecting the application of " looking for a needle in a haystack " formula of the cell of blood extremely low concentration, for example search for the tumour cell that circulates in blood.
The experiments experiment example
Embodiment and operation in order further to illustrate system described herein, apparatus and method, provide the following example.These embodiment are not interpreted as limiting system described herein, apparatus and method, and more should be interpreted as comprising any and all variants, and it becomes apparent as disclosed result provided herein.
In implementing hereinafter the experiments experiment example, use the synthetic cobalt-ferrite nanometer particle of co-precipitation program, finally merge to (Khalafalla SE, Reimers GW (1973) U.S. Patent number 3 in the ferrofluid (20% solids content) based on water, 764,540).By adding the potpourri of cobalt chloride hexahydrate (II) and iron chloride (III), cobalt ferrite nanoparticles is precipitated out from the 1M sodium hydroxide solution of boiling.The magnetic precipitation uses DI water to clean twice.Add 2 M nitric acid and 0.35 M ferric nitrate (III) solution (Massart, 1981, IEEE Trans Magn 17:1247-1248 in precipitation; Fischer etc., 2008, IEEE Int Conlon Nano/Micro Eng and Molecular Syst China, 907-910).Then potpourri stirred 20 minutes at 80 ℃.Then salpeter solution decants and sediment is fixed on appropriate location with magnet.Vectolite particle in precipitation was dispersed in DI water afterwards, and the ferrofluid that obtains is to sodium citrate and citric acid solution (pH level 7.4) 1 week of dialysis of 40mM.Between dialysis period, solution upgraded on basis in every day.The ferrofluid that obtains is 20 ℃ of viscosity with 1.5cP.
In implementing hereinafter the experiments experiment example, use and obtain transmission electron microscope (TEM) image from Tecnai 12 electron microscopes (120 keV) of Philips.With carbon film covering copper/rhodium net (from Electron Microscopy Sciences) and be dipped into ferrofluid sample with ethanol dilution.After obtaining the TEM image, the particle size in image uses ImageJ software to characterize.From the probability density function match of lognormal distribution of TEM image (counting is 200 particles approximately) the magnetic nanoparticle core Size Distribution that obtains, as follows
Wherein
DThe stochastic variable of describing core diameter, and
D 0 Be respectively ln(with σ
D) mean value and standard deviation.
The AC magnetic susceptibility of the ferrofluid of frequency dependence, can change acquisition (Maiorov, 1979, Magnetohydrodynamics 15:135-139) by measuring the right mutual inductance of solenoid in the situation that has and do not exist ferrofluid.In this respect, pick-up loop (200 circles, mean diameter is 9.76 millimeters) concentrated in solenoidal field coil (340 circles, mean diameter are 13.34 millimeters), and the mutual inductance of two coils characterizes by the table of the LCR from Agilent (E4980A).The ferrofluid sample is imported in two groups of coils in the plastic injector of 1cc.Symmetry in arranging guarantees the parallel field line of force in the pick-up loop position and can carry out the mutual inductance analytical calculation according to the data that record, and finally AC magnetic susceptibility carried out analytical calculation.
The magnetization relaxation equation, suppose not have fluid motion or convection current (Rosensweig RE (1997) Ferrohydrodynamics (Dover:New York)), is
Wherein ω is the local vorticity in ferrofluid,
χ 0Be the DC magnetic susceptibility value of ferrofluid, and τ is the relevant magnetic relaxation time constant of magnetic nanoparticle.The uniform magnetic field of cylinder in arranging causes symmetry, and it makes vorticity (and therefore second in above-mentioned equation) can ignore in measurement space.The magnetic relaxation time constant represents the combination of two physics relaxation processes.Magnetic core as fruit granule is enough little, their magnetic moment will rotate simply the inside nano particle (N é el relaxation) (Rosensweig RE (1997) Ferrohydrodynamics (Dover:New York)), and characteristic time constant by
Provide, wherein
f 0That precession frequency is (usually 10
8-10
12In the scope of hertz),
K a Magnetic anisotropy energy density,
V Core The core volume of nano particle, and
k B THeat energy.Particle with larger core will have higher magnetic anisotropy energy, cause the fixing magnetic moment of in-core, and particle itself will be in solution and externally-applied magnetic field rotating Vortex (Blang's relaxation), characteristic time constant by
Provide.
Herein
ηThe dynamic viscosity of fluid,
k B Boltzmann constant,
TAbsolute temperature (Kelvin), and
D hyd Be the fluid mechanics diameter of particle, comprise its surfactant layer.Account for faster the leading position of relaxation process in these two kinds of mechanism.Vectolite has high magnetic anisotropy energy density (to bulk material at 1.8x10
5To 3.0x10
5J/m
3, and for nano particle up to 3.15 * 10
6J/m
3(people such as Tung, 2003,
J Appl Phys93:7486-7488)), and based on the ferrofluid relaxation of this material, be mainly by higher than the about particle rotation of the critical nanoparticle size of 5nm diameter (Blang's mechanism).Greater than this critical dimension, only Blang's time constant is considered in the measurement of the AC magnetic susceptibility of explaining us due to most of nano particle of observing in the TEM picture.
In the situation that there is no vorticity, the sinusoidal steady-state solutions of equation 2 produces the concept of effective susceptibility, and it describes the magnetization and the amplitude in the magnetic field that applies and the relation between phase place of ferrofluid as the function of frequency:
(equation 5)
(Debye?PJW?(1929)?Polar?Molecules.?(Dover:?New?York))。Herein,
χ 0Be the DC magnetic susceptibility value of ferrofluid, and τ is the relevant Blang's relaxation time of magnetic nanoparticle.
Ferrofluid is comprised of the particle with Size Distribution (typically being lognormal distribution), and it also causes the relaxation time to distribute.Consider this point, the linear combination of all magnetic susceptibility spectrums that the particle size that whole AC magnetic susceptibility is described as existing in ferrofluid produces, by the probability density function of the lognormal distribution of given interrelation of particle size
F(D hyd )Metering:
In nano particle, the amplitude of total magnetization intensity is directly proportional to its core volume, and its indivedual contributions to the magnetic susceptibility spectrum are also like this.Therefore, in equation 1 probability density function by
V 2 Core Weigh.Normalized factor A is provided by following formula
(equation 7).
Suppose hydrodynamic diameter lognormal distribution (equation 4-7), AC magnetic susceptibility data (example as shown in Figure 8 B) can be used the sinusoidal steady-state solutions match of magnetic relaxation equation.Simultaneously, again suppose the lognormal distribution of identical hydrodynamic diameter (and granule density is as free parameter), the relative shape of DC magnetization data (as shown in Fig. 8 B illustration) can be used the Langevin equation model.Simultaneously the match well explain experimental result, obtain the average hydrodynamic diameter of 72.5 nm.This value is much larger than the average core diameter of the nano particle that obtains with TEM.Reasonably explain for one of this difference to be, in balance, nano particle forms middle-sized gathering, the magnetic field that applies during measuring as single unit response.Sample to the dilution of same ferrofluid has also carried out dynamic light scattering experiment.Those results have confirmed hydrodynamic diameter much larger than core diameter, support explanation in this paper.
The concentration of the surfactant that typically, uses is high enough to prevent colloidal stability degenerate continuously (at least in some months).Therefore, possible particle aggregate forms at one of brief precipitate phase of ferrofluid synthetic schemes, is speed up processing, and it often relates to the use of permanent magnet.Surfactant added afterwards, can not smash the gathering that has formed.
In the further enforcement of experiments experiment example, use the ZetaPALS equipment from Brookhaven Instruments Group to carry out dynamic light scattering experiment hereinafter.For these measurements, ferrofluid is avoided multiple scattering with the dilution of DI water.Find that the fluid dynamics particle diameter is 64.9nm.
In the extra enforcement of experiments experiment example, the particle operating means that uses in experiment (for example, the device shown in Figure 1A 100) comprises two parts: microchannel and following copper electrode hereinafter.Electrode (30 μ m are high, and the wide and 2cm of 300 μ m is long) can be by use photoresist mask wet etching thermal envelope the copper layer of printed circuit board (on the insulated metal base material) make.Produce by applying with the alternating current of electrode single-layer orthogonal the magnetic field of advancing in passage.Microchannel (20 μ m to 100 μ m are high, and 1mm to 3mm is wide and 2cm to 3cm long) can pass through the soft lithography stamp, from dimethyl silicone polymer (PDMS) preparation, and with the very thin PDMS insulation course of coated electrode, be connected (Mao et al., 2006,
Nanotechnology17:34-47).Selection is far below the channel height of local flowing molten iron body dynamics stream optimal value, in order to minimize it for the potential effect of particle moving.In the separating experiment of the tracer grain that uses submicron order, do not observe recognizable fluid dynamics stream.The insulated metal base material can efficiently radiates heat, and the AC electric current by electrode can be up to 10A under low-voltage thereby make.Before introducing ferrofluid/mixture of microspheres in microfluidic devices, passage cleaned approximately 10 minutes with 1% Triton-X solution, to minimize the particle that is attached on the PDMS wall.
In the enforcement of experiments experiment example hereinafter, from Duke Scientific (Fremont, CA, USA) obtain the green fluorescence polystyrene microsphere of different size (1.2 μ m, 1.9 μ m, 2.2 μ m, 3.1 μ m, 5.0 μ m, 6.0 μ m, 9.9 μ m diameters).The microsphere diameter coefficient of variation is approximately 1%.The microballoon of these customized production has low-down factor of porosity and carries in its surface the charged group of minimum.Microballoon is suspended in deionization (DI) water, and is kept at 4 ℃, until they are used for the ferrofluid experiment.
In addition, for make cell in ferrofluid as seen, haemocyte with green fluorescent membrane dyestuff PKH67(available from Sigma-Aldrich) dyeing.This dyestuff has excitation peak at 490nm, at 502nm, has emission peak (Horan et al., 1989, Nature 340:167 – 168).According to the scheme of manufacturer and there are some modifications to carry out cell dyeing.
The common preparation scheme that is used for carrying out experiments experiment example hereinafter is as follows: get blood from the contributor before experiment, be stored in 4 ℃ before dyeing., with about 1,000 ten thousand cell centrifugations, remove subsequently blood plasma.Then cell is suspended in 500 μ l RPMI 1640 nutrient culture media (available from Invitrogen, Carlsbad, CA, USA) that do not contain serum, and fully mixes to remove any adhering to and the cell of combination.The cell suspension that obtains was 1000rpm centrifugal 5 minutes again.
Carefully blot supernatant, and bead is suspended in 500 μ l dilution C (being supplemented with staining kit).At once afterwards, be prepared in 4 micromole PKH67 dyestuffs in dilution C.Mix isopyknic dyestuff and cell solution.The cell suspension lucifuge that obtains was hatched 4 minutes.By adding equal-volume hyclone (FBS) to stop staining reaction, and cell suspension was further hatched 1 minute.Cell then centrifugal 5 minutes of 1200rpm to remove staining solution.They clean 3 times to remove the dyestuff in any remaining solution in containing the cell culture medium of 10%FBS.After cleaning fully, cell is suspended in nutrient culture media.The brightness of labeled cell is tested with fluorescent microscope.Before mixing ferrofluid, the cell of dyeing Dulbecco phosphate buffered saline (PBS) (PBS) buffer solution for cleaning that contains 10%FBS.
In addition, because citrate is effective surfactant in ferrofluid, and be biocompatible in most of situation in cell is cultivated, so citrate is used as stable ferrofluid, and provides and make cells survival ionic medium wherein.Under this background, determine that debita spissitudo is important because very little or too many citrate will cause particle aggregation and precipitation in ferrofluid.And the survival of ferrofluid inner cell depends on that having enough ionic species promotes continuation with the osmotic pressure of controlling on cell.Still cause the highest citric acid salt concentration in the ferrofluid of the stable soliquid of magnetic nanoparticle to be confirmed as approximately 40mM.Higher citric acid salt concentration will start stabilization removal ferrofluid gradually.
Adopt trypan blue (available from Invitrogen) staining technique monitoring cell viability.Trypan blue is that selectivity dyes dead cell for blueness, dyestuff (The Sigma-Aldrich Handbook of Stains, Dyes that living cells and dead cell can be distinguished; Indicators, Green, F.J., ed., Aldrich Chemical Co. (Milwaukee, WI:1990), 721-722).According to the scheme of manufacturer, adding 10 μ l concentration to 90 μ l 0.4% Trypan Blue liquid is in every 1 milliliter of nutrient culture media 5 * 10
5The cell suspension of individual cell.After at room temperature hatching 5 minutes, the small sample of taking from this potpourri is placed to hemocytometer with living cell counting.The least concentration of having determined at first to cause the citrate (with the stable pH 7.4 that obtains of citric acid) of most cells survival during several hours be 40mM(as shown in Figure 8 C).Under this ion concentration, cell has vigor and ferrofluid to stablize.Therefore, in all experiments of cell, use the citric acid salt concentration of 40mM in relating to the ferrofluid that is suspended in us.
Embodiment 1: ferrofluid character and device characteristic
The concentrated ferrofluid of height that use contains living cells always is proved to be a challenge, because it needs well-designed colloid system.Keeping the maximally related ferrofluid parameter of living cells comprises: pH, ionic strength and nano particle-surfactant combination, and together with them integral body and relative concentration.
It is very important finding this respect that is combined in of suitable nano particle-surfactant: ferrofluid need to be stable at pH 7.4, and colloid-stabilised sexual needs keep up to the ionic strength that can keep living cells.Should also be noted that the Size Distribution of the nano particle in ferrofluid.If there is diameter to only have the nano particle of several nanometers, they can pass through cell membrane, and cause direct cytotoxicity (Scherer etc., 2005, Brazilian J Phys 45:718-727).For this reason, system described herein, apparatus and method comprise the magnetic settling step in synthesizing of biocompatible ferrofluid, specifically the nano particle of minimum is stayed.
The commonsense method of improving the biocompatibility of ferrofluid typically relates to thick polymeric layer, forever cover the magnetic nanoparticle (people such as Bautista as glucosan, 2004, Aranotechnology15:S154-S159), because surfactant molecule reduces toxicity by the surface that hinders direct contact inorganic nanoparticles.Yet such method causes the volume content of the magnetic nanoparticle in ferrofluid significantly to reduce, and the corresponding decline of its magnetic susceptibility.The magnetic susceptibility of higher ferrofluid typically is converted into particle operation faster, so ferrofluid of the present disclosure is by using short surfactant molecule optimization.
In some embodiments, the ferrofluid that uses comprises and suspending in water, and with the cobalt ferrite nanoparticles of citrate-stable.Be found to be approximately 11.3 ± 4.4 nanometers (as shown in Figure 8 A with the average nano particle core diameter in the definite ferrofluid of transmission electron microscope (TEM), show the figure that describes cobalt ferrite nanoparticles Size Distribution in the ferrofluid that obtains by TEM, engineer's scale is 50nm).According to the synchronous match to AC magnetic susceptibility and DC magnetization data, average hydrodynamic diameter is confirmed as approximately 72.5 nm.Especially, as shown in Figure 8 B, show to describe the AC magnetic susceptibility of ferrofluid and the figure of demagnetization curve, the match to AC magnetic susceptibility data of supposing the lognormality Size Distribution shows the moderate particle aggregation of average hydrodynamic diameter 72.5 nm.Difference between the average hydrodynamic diameter of observing in the TEM image and independent core size shows in the soliquid of ferrofluid particle aggregation to a certain degree.This discovery also confirms by dynamic light scattering measurement, obtains the approximately average hydrodynamic diameter of 64.9 nm on its sample of ferrofluid at high dilution.Yet it is still enough little that magnetic nanoparticle and the microballoon of micron-scale are compared with cell, so that ferrofluid is approximate as continuous magnetic medium.
In building-up process, determine that best ion concentration in ferrofluid is about 40mM, to provide good balance between cell viability (by Trypanblau test, determining) and ferrofluid stability.Especially,, with reference to figure 8C, provide the figure that describes viable count and citric acid salt concentration.As shown in the figure, the combination that is considered to cell viability and ferrofluid stability of the citric acid salt concentration of 40mM (stable to obtain pH 7.4 with citric acid) is best.Dotted line is illustrated in the cell count in original blood sample.Count 3 correspondences and consume the cell of ≈ 1h in citrate solution.In the process of given experiment, cell keeps its vigor.Observe 75% cell and maintain vigor,, even after suspending several hours in ferrofluid, make to relate to the living cells operation and can be extended with the test that separates.
Before the cell manipulation experiment, ferromagnetic microfluidic device uses fluorescence polystyrene microsphere (Duke Scientific; Single scattering device, diameter range is 1.2 to 9.9 μ m)., in order to understand the impact on the behavior that is dispersed in the non-magnetic particles in ferrofluid of excitation frequency and current amplitude, use the microballoon of different size to carry out series of experiments under different excitation frequencies and current amplitude.The microballoon mixing ferrofluid of a small amount of intended size (for the mean particle dia of minimum, every milliliter at the most 1.1 * 10
6Individual microballoon), and subsequently add microchannel.Feeder connection and outlet two ends clamp, to prevent the transient state fluid motion.Near the microchannel top microballoon uses upright fluorescent microscope (Zeiss Axiolmager A1) and highly sensitive video camera (Retiga2000R) (adopting StreamPix software) from upper surface imaging.At MATLAB(MathWorks) in carry out the offline image analysis by the light stream program.This program can be in the visual field in less than one (1) minute pursuit path automatically, and determine the size of thousands of independent microballoons.
In these experiments, the dynamics of having observed two class particles.When low frequency, microballoon is positioned between electrode, and wherein the repulsive force that causes of magnetic field gradient forms local minimum.With reference to Fig. 9 A, higher than critical value
f c Frequency, cause microballoon along the continuous translation of channel roof length.This threshold frequency depends on particle size and electrode separation, but must not depend on the amplitude (Fig. 9 B) of input current.The average velocity of intended size microballoon depends on the position of amplitude with their relative lower electrodes of excitation frequency, electric current.
Especially, Fig. 9 A and 9B show the function of particle speed as incoming frequency and current amplitude.In Fig. 9 A, middle figure is presented under the input current amplitude (peak-peak) of 7-A, under two different frequencies, and the space distribution of the instantaneous average x speed of diameter 6pm particle.Due to the repulsive force from magnetic field gradient, particulate slows down or stops fully between electrode.The equilibrium point (being particle capture) that the zero passage negative slope is corresponding stable.The figure demonstration at Fig. 9 A top, at 10 Hz, particle trajectories stops between electrode, causes and catches.The figure demonstration of Fig. 9 A bottom, at 4,640 Hz, particle is continuous moving in passage length.This comes the leading zone that surpasses repulsive force of magnetic moment of the local rotational component of ripple voluntarily.The stain of every track end represents the final stopping place of particle.Fig. 9 B show higher than threshold frequency (
f c ) after, the microballoon of 6 μ m rolls continuously and is not hunted down along the top passageway surface.
The dependent particle separation of Figure 10 A-10D display frequency.In the experiment of various microsphere diameters, finding increases with particle size, and the threshold frequency monotone increasing shows by excitation frequency and controls potentiality for the particle separation based on size.Figure 10 A demonstration threshold frequency (
f c ) the particle size dependence.The particle of different-diameter is discrete
f c Value makes by being transferred to suitable frequency can carry out separation based on size.This phenomenon can be explained by the reasoning of simple hydrodynamic force mechanics.Magnetic force and magnetic moment particle volume (
R 3 ) weigh, the hydrodynamic force mechanical resistance that resists the linear particle motion makes with relative magnetic moment that particle rolls with the R of relative magnetic force
R 2 Weigh.Therefore, the linear particle speed that causes separately of magnetic force depends on
R 2 , and magnetic moment causes, with R, weigh.This observation shows, magnetic moment is relatively more remarkable on the impact of less particle, and has explained why less particulate can overcome the repulsion that magnetic force is caught, and diffusion continuously in passage under low frequency more.Block curve shown in Figure 10 A represent the analog result of threshold frequency and well explain nanometer (1 nanometer) average microballoon wall gap and be applied to the data of non-slip condition of the rotation of microballoon.
Figure 10 B shows the average velocity (also referred to as operating speed) of microballoon (ratio in 8:1 in identical ferrofluid is mixed) under the excitation frequency of 10Hz to 100kHz of 2.2 and 9.9 μ m.As shown in Figure 10 B, 2.2-can separate with 9.9-μ m particle under 400 Hz.For wide frequency range, less particle shifts continuously, and larger particle is trapped between electrode.In specific and other experiment, particle/cell mixture finally is divided into two groups at this, and for example, those captive relative those are removed from passage.Hypothetical target particle/cell is that those intentions are used for catching, and capture rate can be defined as the ratio of the number in its corresponding original mixture of number of the target part in captive group.Equally, separation efficiency is defined as the number of non-target part in the group that is eliminated and the ratio of their corresponding numbers in original mixture.Yet, particle/cell purity be only the number of catching group internal object cell with this group in the ratio of total cellular score.Under the excitation frequency of 400 Hz, the microballoon (167 in 173) of 96.5% 9.9-μ m was hunted down in 10 seconds, and 2.2-μ m particle (1, in 294 1,285) continue to shift and to be eliminated out form (45s) and be not hunted down (as shown in Figure 10 C and 10D) has 99.3% separation efficiency along passage.Catching particle purity in group is 94.9%(176 167 targets in the particle that always is hunted down).The most minimicrosphere of failing to remove from passage is bonded at random position on dimethyl silicone polymer (PDMS) wall, rather than is hunted down between electrode.It is better that passage prepares, and separation efficiency and particle purity can be higher.
Figure 10 C contains the fluorescence microscopy images of a section of the microchannel of 2.2 and the 9.9-μ m microballoon of random dispersion in passage before excitation just.Perpendicular line represents boundary electrode.Figure 10 D is that (the 6A peak-peak 400Hz) is opened the passage snapshot of rear 45 seconds in excitation from position identical shown in Figure 10 C.9.9-the particle of μ m is located in spacing fast between nearest electrode, and the microballoon of 97% 2.2-μ m is advanced continuously from right to left and is not hunted down.In Figure 10 D in the visual field nearly all less microballoon as new, a collection ofly from the right, enter.
Figure 11 A and Figure 11 B describe the cell separation of bacterium and haemocyte.Figure 11 A be described in comprise the Escherichia coli bacterium (
E. coli) and erythrocytic sample under 200Hz the space distribution of x speed.Under this frequency, most of red blood cell is hunted down (the zero local velocity by them shows) between electrode, and Escherichia coli can be slowly but move through continuously this zone.As described herein, the fluctuation in the RBC number certificate is statistical property.Figure 11 B describes the sickle cell and separates.The sickle cell, compare shape with elongation and the elasticity of change with normocyte, be hunted down between electrode and concentrate, and healthy cell still can circulate under 300Hz in microchannel.The electrode separation of the device in Figure 11 A is different from Figure 11 B, causes in each passage erythrocytic
f c Different.
Figure 12 A describes the average velocity of particulate, by the maximal value of square relative 12A of current amplitude (peak-peak), carries out normalization, is described as the function of excitation frequency.Square being directly proportional of particle speed and electric current, until about 7A.Figure 12 B is described under the 6A peak-peak, the mean speed relative frequency that the 9.9pm microballoon is advanced at the electrode of different spacing.Less spacing causes higher particle speed and less threshold frequency.Figure 12 C is described in the concept sketches of the particulate sorter on the basis of the effect of observing in Figure 12 C.Under given excitation frequency, less spacing is caught larger particle, and allows less particle pass through.Finally, even minimum particle also can be trapped in larger gap.Therefore, suppose that particle moves from left to right, and the passage of described electrode is acellular at first.
Therefore, movement of particles is confirmed as depending on electrode separation, and less spacing causes microballoon faster to be advanced and the minimizing (referring to Figure 12 B) of threshold frequency.This phenomenon can be used for device, and the electrode zone that it is characterized in that having different gap separates the granulate mixture that has more than two different sizes with the excitation frequency with same.Electrode mode can also use gradually the gap that increases to create, with based on size separation particle (for example, as shown in Figure 12 C).In this respect, observe the medium and small unevenness (manufacturing causes) of actual electrode separation and partly determine the resolution of separating, the size difference of minimum in the particle that being defined as still can be efficiently (for example,〉90%) separates.The particle size of given certain limit, this isolation resolution is directly involved in the difference in corresponding threshold frequency.(that is, preferred electrode gap and the very rare cell concentration of controlling) under ideal conditions, isolation resolution can be arbitrarily small.But around each inhomogeneous electrode gap, threshold frequency tends to show slight localized variation.As shown in Figure 10 A, best threshold frequency non-linearly depends on particle radius.Therefore, between 9-μ m and 10-μ m microballoon, the 1-μ m diversity ratio 1-μ m of diameter and the intergranular 1-μ of 2-μ m m difference are more easily differentiated (slightly random variation is arranged in electrode separation).The resolution of the separation that finally, realizes in experiment described herein is ≈ 1 μ m for 2 μ m or larger particle.
Embodiment 2: the impact of current amplitude on particle speed
In additional experiment, study and determined the dependence of particle manipulating speed as input current amplitude function.According to the calculating of general introduction herein, and the supposition ferrofluid keeps linear magnetic, the particle speed square measurement of input current.As shown in Figure 12 A, this hypothesis starts to break through the peak-peak input current amplitude higher than 7A.
Electrode separation also changes (electrode width is fixed on 210 μ m) to determine its impact on the particle operation.Less electrode separation causes average grain speed and lower threshold frequency (as shown in Figure 12 B) faster.This observation can be explained the local magnetic field gradient that particulate produces magnetic force by pointing out the more approaching minimizing of electrode separation.More approaching electrode is also incited somebody to action more multi-energy and is put into the fundamental frequency that these particulates is produced the capable ripple of magnetic moments.Lower magnetic force and the magnetic moment of Geng Gao cause particulate rotation faster and whole gait of march.They also cause threshold frequency lower (Tung LD, et al. (2003) Magnetic properties of ultrafine cobalt ferrite particles.
J Appl Phys93:7486-7488).This observes the idea of further supporting to use granule separator, and wherein the spacing between electrode can increase to catch more and more less particle or cell (as shown in Figure 12 C) gradually.
Embodiment 3: the separation of living cells
Based on the physical behavio(u)r of ferromagnetic miniflow platform, with the human red blood cell of living with bacterium operates and separating experiment, to prove effectiveness and the practicality of ferromagnetic micro-fluidic system and device for biomedical applications as herein described.Red blood cell and Escherichia coli bacterium [the K12 bacterial strain (Blattner etc., 1997,
Science277:1453-1474))] dyeed and mixed with green fluorescent marker before being suspended in ferrofluid.Cell in passage and the average velocity of bacterium are measured under the frequency from 10Hz to 100kHz with the peak-to-peak current amplitude of 6A.The threshold frequency of finding cell and bacterium is respectively 215 and 77Hz.These
f c Value is slightly lower than the suitable polystyrene microsphere of size, may be due to the shape of complying with and causes the non-spherical geometric combination that increases along the difficulty of channel roof rolling.In addition, have bacterium and the cell of its complicated surface chemistry, with the interaction of PDMS passage,, than naked microballoon stronger (causing more general cell to adhere to), show higher effective kinetic friction coefficient potential between cell and PDMS surface.Refer again to Figure 11 A, shown under the excitation frequency of 200 Hz that cell and bacterium are along the linear velocity of the space average of passage.Less Escherichia coli are moved (speed point in Figure 11 A does not have zero passage) continuously along passage, and finally leave form, and haemocyte is positioned (speed point reaches zero) between electrode.Should be noted that, the larger variation of observing in the RBC number certificate of Figure 11 A comes from the fluctuation of statistics: only have a small amount of red blood cell to pass through given x position in form, and their non-spherical form means, when it rolls passage in this position, each cell will be random angular orientation (with slightly different instantaneous velocity).Bacterium,, although length is different and non-spherical, also have enough numbers (hundreds of passes through given x position) to obtain good average statistics.Finally, 6,750 of ≈ that are presented at first in 7,050 Escherichia coli in the sample visual field were eliminated (95.7% separation efficiency) in 45 seconds.1,018 erythrocytic 954 that present at first are hunted down, corresponding 93.7% capture rate and 76.1% cell purity.
In different experiments, healthy red blood cell and those suffer from sicklemic cell separation, and this is by utilizing shape and the flexible difference (as shown in Figure 11 B) between them.To comprise ratio approximately the health of 4:1 the erythrocytic blood sample of falciform is added to ferrofluid and imports microchannel.At 300Hz, the sickle cell is hunted down, and healthy haemocyte is eliminated (fluctuation of each data centralization shown in Figure 11 B is statistical property) continuously from passage.In the red blood cell that comprises at first 501 health of ≈ and 145 drepanocytic samples, 300 healthy cells are eliminated, and 109 sickle cells are hunted down.Hypothetical target is to remove sample from the sickle cell, separation efficiencies of these numerals corresponding 75.2% (109 in 145 sickle cells are separated from healthy cell), and 89.3% healthy cell purity (300 healthy cells and 36 sickle cells are eliminated).
The ferromagnetic microfluid of these examples proves is delivered to sensor array by quick separation with the target cell selectivity in raji cell assay Raji, significantly reduce incubation time and increase the purposes of diagnostic sensitivity.Although in microfluidic devices the operation of particulate and living cells with separate also may be by having set up technology (for example DEP and based on the method for magnetic mark) carry out, ferromagnetic microfluidic methods as herein described has many advantages compared with the conventional method.For example, target cell can be effectively, rapidly, and in unmarked mode, be concentrated, catch, locate or orientation sensor surface simply.Biocompatible ferrofluid of the present disclosure can make the cell of invigorating blood circulation keep several hours, and physical property is not degenerated, and allows to extend the detection of target sample.
When simple photodiode is combined, the ferromagnetic microfluidic separation of cell can provide fast, robotization and disposable blood measure, it can, in the concentration of for example inside counting in 1 minute and any target cell type of estimation (as bacterium or sickle cell), not need microscope, pump or tediously long sample preparation steps.System as herein described, apparatus and method can also be used for the concentrated rare cell of selectivity, the circulating tumor cell in blood sample for example, difference (Lekka et al., 1999 of the Young modulus (Young's modulus) by utilizing the target cell type
Eur Biophys J28:312-316).System as herein described, the apparatus and method of application can increase the detection sensitivity of existing raji cell assay Raji in this way.
Thereby system as herein described, apparatus and method are included in interior cell manipulation and the separation platform that uses biocompatible ferrofluid of microfluidic devices cheaply.Proved and can realize that high-efficient granule separates being less than in one (1) minute.As an example, bacterium can separate from the haemocyte of living, and the sickle cell can separate from healthy red blood.For the device based on stream, separation can realize by the particle operation vertical with flow path direction.By geometry and the input excitation frequency that changes electrode, system as herein described, apparatus and method are applicable to particle and the cell of different size scope.Together with the control of the surface chemistry of microchannel, system as herein described, apparatus and method can be incorporated in chip lab sensor and diagnostic system with active region that target cell is led.In this mode, system as herein described, apparatus and method can reduce significantly incubation time and increase in existing sensor and diagnostic platform the actual detection sensitivity that realizes.
2. target substance is concentrated
As described, this paper goes back descriptive system, device, method and other embodiment, comprise and concentrate the device that is suspended at least a target substance in biocompatible ferrofluid, it comprises and is configured to accept to comprise the sample of at least a target substance and the microchannel of biocompatible ferrofluid, and at least a target substance of being accepted in sample is concentrated in the inlet flow with correlated inputs width substantially.This device further comprises at least two electrodes of the contiguous microchannel in position, described at least two electrodes are configured at least when the controllable electric electric current is applied to described two electrodes, produce controlled magnetic force in containing the sample of ferrofluid, the controllable magnetic that produces causes at least a target substance to concentrate in the stream that obtains, and its width is narrower than the relevant input width of inlet flow.In some embodiments, the magnetic force that produces can apply to control by the electric current at least to electrode controlled (that is, with about controlling the described similar fashion in magnetic field that is used for making the discontinuous separation of plurality of target substance).
As described herein, the advantage of microchannel and electrode technology is that it concentrates the ability of with concentrating cells, arranging in single file in channel roof.Therefore,, due to the electrode of potential tiny model, have this ability and can realize high precision.Use is from the magnetic field of external permanent magnet, also can realize concentrating to a certain degree.Cell is pushed to the top of passage, and concentrated them to make it be easy to detect, count and catch them.
As will be described in more detail below, at least three kinds of modes are arranged, wherein can use Different electrodes geometry to realize that cell concentrates in ferromagnetic microfluidic devices: at least two parallel poles that (i) along the length of passage, carry DC or AC electric current (with different relative phases) are shifted cell onto in the space of the top, gap between them; (ii) at least two groups are in the opposite direction carried the electrode in the magnetic field of advancing, and can make cell roll to border between every group; (ⅲ) carry DC or AC electric current (at different relative phases, comprise those that cause advancing), the array of the electrode that adjacent gap reduces gradually, will promote and cell will be remained in those gaps, until that gap becomes is enough little, so that the effect of magnetic moment becomes outstanding.Front two kinds of methods can be used for target cell stream is concentrated near detecting device/counter, and the desirable large-scale parallel that is used for realizing the input sample of the third method is concentrated and separating subsequently.Particularly, in the third method, at first target substance (for example, cell) can be concentrated in alternately and catch in gap, and target cell will be split in adjacent segment subsequently.
The advantage of using electrode and microchannel Configuration cell to concentrate is as follows:
Because there is no fluid dynamics sheath stream, can realize very effective concentrating, the whole volume of device can be the special use of input stream of cells.Therefore, use embodiment described herein to compare the existing method of using fluid dynamics to concentrate and can realize more high flux.
Focus on electrode gap every one by inputting stream of cells, target substance (for example, cell or pathogen) can be separated to adjacent gap.This ability makes in separation and sorting function and has realized theatrical walking abreast, because the characteristic of this equipment is to cross over many such electrode mode that the circulation road width is arranged.
When lining up single file near the top of cell at passage, they can be detected (, by variety of way, comprise, for example, light, magnetic, electrical impedance and capacitance measurement) and counting at an easy rate.By this way, the systems/devices that realizes can be used as " nano cell counter " and uses, and can separate and count millions of cells within a few minutes.Systems/devices can, as the detecting device of pathogen, maybe can be used for helping to characterize the concentration of other non-target cells (as CD4, red blood cell, blood platelet etc.).
With reference to Figure 13, show the schematic diagram of microfluidic devices 500, it comprises the electrode group, it is configured to produce magnetic field and focuses on basic single file stream with the target substance with sample.Device 500 comprises microchannel device 510 and electrode group 520.Microchannel 510 can be similar to Figure 1A, 4 and 5 described microchannels.Electrode group 520 comprises, in the example embodiment of Figure 13, comprises two " wavy " electrodes parallel to each other basically usually.Produce magnetic field while to the electrode group, applying controlled at least a electric current, the target substance that causes being suspended in ferrofluid is pushed and concentrates in microchannel.The controlled at least a electric current that is applied to electrode can use controller for example the controller 160(of Figure 1A can be based on the controller of processor) control/regulate.
In some embodiments, described at least two electrodes (for example electrode 520) can comprise having following at least a structure, for example, one or more straight shapes basically of at least two electrodes, one or more corrugated shape basically of at least two electrodes, the substantially parallel layout of at least two electrodes, and/or at least two electrodes location of taper basically, wherein said at least two electrodes are near each other gradually.
In some embodiments, the electrode group is configured to produce magnetic field, and when applying described at least a electric current thereon the time, it is applied to target substance at the entrance area of microfluidic devices basically.In these embodiments, a plurality of electrode groups can be used to microfluidic devices, do in order to concentrate the sample that comprises at least a target substance in the single file arrangement of the microchannel of flowing through for one group, and another electrode group, its fundamental purpose is (for example to implement discontinuous separation, to the continuous stream based on ferrofluid) sample, separating two or more different target substances to relate to for example described mode in Fig. 1-7.
In some embodiments, concentrate on the magnetic force of microfluidic devices entrance, can cause with cell be sorted into fast define in microfluidic devices catch gap (for example, in the passage of this device).Therefore, this centralized function can be used at first concentrating target substance (for example, target cell) to alternately catching, and then target cell is separated to adjacent segment.
Therefore,, with reference to Figure 14, show and be configured to target substance to be focused on the alternately schematic diagram of the part of another example microfluidic devices 600 in gap.Device 600 comprises microchannel 610, and it can be similar to about Figure 1A, 4,5 and 13 described microchannels.Device 600 further comprises one group of electrode with a plurality of electrodes 620, and it comprises at least two class geometric electrodes.In the example of Figure 14, one (definition pair of electrodes) of every two adjacent electrodes has the length of the entrance area 630 that basically extends to device, thereby with definition set electrode 622.Adjacent with each this concentrated electrode 622 is shorter electrode 624.In some embodiments, can use pair of electrodes, it comprises concentrated electrode 622 and shorter electrode 624, and these arrangement of electrodes are the location of taper basically relative to each other, thereby they are near each other.A plurality of electrode 620(comprise concentrate electrode and shorter electrode) from microchannel one lights has wavy shape, it is through entrance stage, and extends to the exit region that installs.In some embodiments, from the locus that a plurality of electrodes 620 are taked common wavy shape, electrode extends with direction basically parallel to each other usually.
Example geometry according to electrode shown in Figure 14, while to a plurality of electrodes 620, applying at least a electric current, the geometry of the electrode around entrance area causes producing magnetic field, this causes target substance to be directed to a plurality of single file streams, and it concentrates on alternately catching in gap of electrode pair definition and (comprises the gap 640 of catching shown in Figure 14).In case the stream that flows (its each can comprise the similar mixtures of the target substance that device portal is accepted) forms, the magnetic field that acts on a plurality of sample flow of concentrating now causes different target substance separation to enter adjacent segment (namely,, as the result of centralized operation, do not accept the gap of any target substance).Magnetic field around the magnetic field (this place's electrode 620 is parallel basically, and it operates to carry out discontinuous lock out operation) that the flow channel area place forms is compared in inlet region can have the attribute of different (or similar).The discontinuous lock out operation that is undertaken by the magnetic field of substantially parallel electrode 620 positions is similar to the operation of carrying out about the described system in Fig. 1-12, equipment and method.
Therefore, in some embodiments, device can comprise electrod-array, and at least some of array the first arrangement of electrodes, for become the location of basic taper with respect to adjacent electrode, makes described the first electrode be configured to move closer to adjacent electrode.Electrod-array be configured to produce stream that magnetic force causes at least a target substance that obtains adjacent electrode right above and between form.Thereby the centralized operation rear electrode between each gap become gradually empty, but when flow stream carries out lock out operation, empty gap acceptance flows in adjacent segment at least a target substance that separates from potpourri, wherein said potpourri is concentrated in adjacent segment.
Learn and can be used for exercising centralized function from those other different geometric electrodes shown in Figure 13 and 14.
In some embodiments, the electrode that is used for carrying out centralized operation can be configured to produce at least two magnetic waves of backward-travelling wave field, to cause at least a target substance, focuses between the magnetic wave of at least two generations near the center, border., with reference to Figure 15, shown the schematic diagram of part of another example embodiment of microfluidic devices 700.Device 700 comprises microchannel 710, and it can be similar to about Figure 1A, 4,5,13 and/or 14 described microchannels.In the embodiment of Figure 15, device 700 further comprises one group of electrode with a plurality of electrodes 720, has common wavy shape, and has relative to each other substantially parallel location.The contiguous microchannel 710 in electrode 720 positions.For example, electrode 720 can be arranged in the electrode layer that is arranged in the microchannel outside.
While to the electrode of Figure 15, applying controlled at least a electric current, produce two oppositely row magnetic waves (being labeled as ripple 730 and 732).Oppositely two magnetic waves of row cause target substance in sample flow to be concentrated near border 740 between the magnetic wave of two generations (being described as dotted line).
The configuration of other electrode can be for generation of the magnetic field (thereby with produce magnetic force) that is used for concentrating at sample target substance.In some embodiments, be configured to produce at least two electrodes be used to the magnetic field of carrying out centralized function and can conduct the controlled AC electric current with the controlled attribute that is defined as implementing desirable centralized function.In some embodiments, at least two these electrodes can the conduct direct current electric current.In some embodiments, single electrode can wrap up self to form two (or more) parallel arms, and it defines a plurality of parts of electrode for generation of centered magnetic field.In some embodiments, one or more permanent magnet groups can be used for supplement or alternative electrode group to help the enforcement of centralized function.
, with reference to Figure 16, shown the process flow diagram of concentrating the example procedure 800 of target substance in microchannel.Program 800 comprises that accepting 810 comprises the sample that is suspended at least a target substance in biocompatible ferrofluid, and at least a target substance in the sample of acceptance is concentrated in the inlet flow with relevant input width substantially.
After accepting sample, at least a electric current (for example, AC electric current with controlled frequency, phase place and/or amplitude, DC electric current etc.) controllably apply on the electrode of the contiguous microfluid in 820 at least two positions, to produce controlled magnetic force in comprising the ferrofluid passage of sample.Magnetic force causes at least a target substance to focus in the stream that produces, and its width is narrower than the relevant input width of inlet flow.In some embodiments, at least two electrodes are configured to conduct the electric current that controllably provides and produce controlled magnetic force, according to, the attribute of the attribute of the attribute of the electric current that applies, microchannel and/or at least two electrodes (for example its structure or shape) for example.
3. the magnetic force of detecting device leads target substance
as mentioned previously, this paper has also described device, system, method and embodiment, the device that comprises at least a target substance in test sample, described device comprises the microchannel of the sample that is configured to accept the biocompatible ferrofluid that comprises at least a target substance and wherein suspend at least a target substance, determine the detecting device of at least a target substance in sample, at least two electrodes with the contiguous microchannel in position, described electrode is configured to comprise when controlled at least a electric current is applied at least two electrodes in the sample of ferrofluid and produces controlled magnetic force.The controlled magnetic force that produces causes at least a target substance guiding detecting device.Depend at least in part the vertical range of target substance to electrode because detect effect and the quality of target substance, if vertical range can not be controlled, dissimilar target substance can not be distinguished reliably.Correspondingly,, by target substance being shifted onto near top (can arrange testing mechanism),, as the result of target substance to the vertical range of detecting device, hinder the challenge that successfully detects target substance and can be controlled and be overcome to a certain extent.
In some embodiments, detection scheme as herein described relates to the direct impedance bioelectrical measurement by the plane electrode of pair of parallel, and/or capacitance measurement.About the impedance measurement detecting device, the cell size scope that need to be counted and detect determines the spacing between these electrodes.Typically, this spacing should be equivalent to or be slightly larger than cell dia.For haemocyte, spacing can be the 10-20 micron.For Bacteria Detection, spacing can be the 5-10 micron.
The existence counting of target substance in sample and the various technique/method that detect are comprised by measure the conductivity of the buffer solution that flows in microchannel through a small amount of electric current between two electrodes.Any fast peak in this electric current (AC or DC) will show that the of short duration variation of the local conductivity of medium-most possibly the process by cell causes, its electric conductivity is different from medium.When the gap between sensing electrode was suitable with cell size, the interference of the electric field patterns between electrode was maximum, and electric signal to noise ratio (S/N ratio) is best.
In some embodiments, can be undertaken by measuring electric capacity detection and the counting of the cell that exists in ferrofluid solution.The electrode lay-out of capacitive sensor is identical with impedance method with cell detection mechanism: pair of electrodes has the width gap suitable with interested cell dia.The electric capacity of measuring is typically little, and it is measured and generally with high-frequency AC excitation (typically,〉1 MHz), carries out obtaining capacitance signal.If detected target substance is bacterium, the relative dielectric constant of these bacterium target substances is the magnitudes 100, and ferrofluid will be in 80 magnitude, so, their existence in ferrofluid can detect, if cell can be positioned at the position near sensing electrode.
Therefore,, by using the magnetic field that produces, for example, use and about described those the similar electrodes in Fig. 1-16, configure, target substance can be pushed channel roof to, and sensing electrode can be positioned herein.In some embodiments, the detecting device that uses in conjunction with microfluidic devices and system comprises antibody " blanket " (target acquisition cell specifically), and a pair of capacitance detector---catching the blanket upstream for one, another is in downstream.In such configuration, sensing capacitor can count and enter (
N_in) and leave (
N_out) capture region cell number and according to calculate poor (that is,
N_in-N_out) define and how much by blanket, caught.Alternatively, can use the electrode pair of measuring impedance, or some other devices of some other character of the number of measurement expression respective intended material/amount.Implementing micro-fluidic system as herein described and equipment is enough strong with the shear flow that obtains on antibody blanket surface, adheres to prevent the non-specific of other cell, to such an extent as to but be not can not cause the loss of target cell too by force.In this operator scheme, different capture regions can be placed on downstream each other continuously, and be a plurality of target substances, for example, pathogen or other target cell such as CD4+ white blood corpuscle, the concrete cancer cell etc. of expressing epithelial cell adhesion molecule (EpCAM) provide quantitative and concrete detectability.
The antibody blanket can be learned and realize by various functionalisation of surfaces.Sensing electrode can be that print, the pattern transfer printing or micro-machined (in evaporation or the sputter of the lip-deep metal of photoetching pattern).
Some advantages of sensor-based microchannel comprise:
By being separated with actual cell sensor (detecting device), functionalisation of surfaces simplifies manufacture process.
The biology sensor direct function sensor surface of a lot of common existing electron devices with integrating.According to micron or nanoscale sensor (it is said and provide much higher sensitivity), functionalized total surface area is very little.Sometimes, only there is a few cell also to have living space to being no more than on sensor surface.Therefore,, if target cell concentration is higher, can guarantee the saturated fast of sensor signal.In order to address this problem, some common devices are arranged to wide array (nearly thousands of) with its sensor, cause the complicacy of system and the challenge of manufacturability significantly to increase.In the system based on miniflow described herein and device, use the detecting device of these systems and device can detect lacking to individual cells through surveyed area; The maximum number of the saturated front cell that can be detected of signal is provided by the relative extent of catching blanket.The dynamic range that increases the sensor that uses in disclosed micro-fluidic system and device can be simply to the scope that increases capture region, and does not need to increase the number of sensor.
Detection method as herein described is unmarked (that is, without fluorescent marker or other mark, is attached on cell and allows to detect).
The use of stream of cells being concentrated and pushed to the current-carrying electrode of surveyed area below passage (or the use of some other magnetic field generation mechanism, for example, mechanism according to permanent magnet), can use input pressure sampling stream, in certain embodiments, less than 1 psi(pound per square inch), keep simultaneously good flux.On the contrary, in common cell manipulation, must use and cell dia (typically 50 microns or less) suitable height and the passage of width, thereby and or the flux of those devices lower, perhaps must use very large source pressure to promote Rapid Flow by these small passages.
Therefore,, with reference to Figure 17, shown the schematic diagram of part of an example embodiment of microfluidic devices 900.Device 900 comprises microchannel 910, and it can be similar to, for example about Figure 1A, and 4,5,13 and/or 14 described microchannels.Device 900 further comprises one or more detecting device 920a-n, one side of position adjacent channel 910 is (in the embodiment of describing, detecting device is positioned at the outer wall of passage 910, the top of its define channel), be configured to detect and microchannel 910 samples of determining to flow through in existence and the number of target substance, and determine type or the identity of the target substance that detects.Device 900 also comprises that the magnetic field generation mechanism pushes the target substance in the ferrofluid sample to microchannel 910 tops to produce, thereby and pushes the magnetic field of detecting device 920a-n to.In some embodiments, the magnetic field generation mechanism can use one group of electrode to implement, for example shown in electrode 930(dotted line), its can be similar for example about shown in Fig. 1-17 and described those.These electrodes can have predetermined configuration, make and produce controlled magnetic field when at least one controlled electric current passes through these electrodes, it pushes nonmagnetic target substance (cell, bacterium etc.) to surperficial with the surperficial relative microchannel near electrode.
For example, electrode can comprise at least two electrodes of the contiguous microchannel in position, it is configured to conduct at least a electric current that controllably provides and produces controlled magnetic force, and this is at least in part according to the physical attribute of electrode, the physical attribute of microchannel and/or the attribute of the electric current that applies.These physical attributes can comprise the structure of at least two electrodes, the structure of wherein said at least two electrodes can comprise following one or more, for example, the substantially parallel layout of one or more corrugated shape, at least two electrodes basically of one or more straight shapes basically of at least two electrodes, at least two electrodes and/or at least two electrodes be the location of taper basically, and wherein said at least two electrodes are near each other gradually.Electrode is configured to work as at least a controlled electric current, and (it can use controller, for example the controller 160 of Figure 1A is controlled) while being applied at least two electrodes, produce controlled magnetic force in comprising the sample of ferrofluid, described electric current comprises association attributes (for example, phase place, frequency, amplitude etc.).
In some embodiments, can use permanent magnet to supplement or alternative electrode, to promote that producing magnetic field pushes target substance to from detecting device 920a-n nearest channel surface.
As described, and as shown in figure 17, can use in some embodiments detecting device based on electric capacity, based on detecting device of impedance etc.For example, detecting device 920a comprises upstream capacitance detector 922a, comprise two sensing electrode plates, capture region (also referred to as the blanket district) 924a, with the downstream capacitance detector 926a that is positioned at capacitor 922a downstream and blanket 924(downstream capacitance detector 926a also as the upstream capacitance detector, be labeled as 922n), be used for the detecting device that the next continuous microfluidic devices in conjunction with Figure 17 900 uses.
By upstream capacitance detector 922a determine through the target substance number after, target substance flows through near antibody blanket 924a, it functionalised to catch, thereby and detects the target substance (cell or some pathogen) of particular type.Blanket 924 helps to obtain specificity in the detection of target substance, and target substance can have similar size and structure (this makes range estimation identify, for example uses microscope, and is more difficult and uncertain).For example, Escherichia coli and Salmonella are shaft-like, have similar Size Distribution, but these target substances can use for the antibody of functionalized one or more antibody blankets and distinguished.
As described, the use of magnetic field generation mechanism (electrode or permanent magnet) can be pushed target substance to enough approach with blanket 924a, to realize more effective capture ability.Subsequently, the target substance of not catching continues to flow and through downstream capacitance detector 926a, and it measures the electric capacity that target substance that residue do not catch causes (or alternatively, impedance), then based on the determined electric capacity of downstream capacitance detector 926a, can determine to remain target substance number (
N_out) (for example, being tested and appraised unit 940).
N_inWith
N_outThereby difference can obtain the number of the target substance of by blanket 924a, being caught.In some embodiments, there is target substance in the determining to show in sample of indivedual members of institute's capturing target substance, if
N_inWith
N_outDifference surpasses predetermined threshold value.Also can use the testing mechanism (for example, based on the testing mechanism of impedance) of other type, determine similarly quantity and the identity of all types of target material in the sample that flows.
In some embodiments, can use the series connection target cell detecting device of similar (or even not same) configuration.For example, in embodiment shown in Figure 17, continuous detecting device 920b in succession after the first detecting device 920a, its blanket district (that is, capture region) can be configured (by suitable functionalized) and catch, thereby and detect dissimilar target substance.
Therefore, in some embodiments, the detecting device that microfluidic devices as herein described and system are used can comprise the electrode at a pair of interval separately, to measure the electric capacity (or impedance) in microchannel,, with the evaluation unit, with the electric capacity based on measured, determine existing of at least a target substance.Identify that variation that unit can be configured to the electric capacity in the measured microchannel that causes based on the existence due at least a target substance determines the existence of at least a target substance.In some embodiments, detecting device can be configured to measure other character that existence affected by target cell, such as impedance etc.Each detecting device may further include capture region, it comprises and being configured to and the interactional material of one of plurality of target material, capture region is positioned at the microchannel downstream of the electrode pair at interval, and another electrode to interval is arranged in the downstream of capture region to measure the electric capacity (or impedance) of microchannel.
As described, in some embodiments, identify that cell location is to determine the initial number of at least a target substance in the electrode pair at interval, with determine the end number of at least a target substance at another place of electrode to interval, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
As further described, in some embodiments, detecting device comprises the test set that the order of series connection arranges, the test set of the order setting of each series connection comprises that the electrode at first pair of interval is to measure the electric capacity (or impedance) in microchannel, capture region comprises and being configured to and the interactional material of one of plurality of target material, and capture region is positioned at the downstream of the electrode at first pair of interval.The test set of the order setting of each series connection also comprises that the electrode at the second pair of interval that is positioned at the capture region downstream is to measure the electric capacity (or impedance) of microchannel, with identify that unit is to determine the initial number of at least a target substance at the electrode place at first pair of interval at each test set place, with the end number of at least a target substance at the electrode place at second pair of interval, and at least partly based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number, whether exceed predetermined threshold value.
, with reference to Figure 19, shown the figure of the example procedure 1000 of at least a target substance in the test sample.Method comprises: 1010 microchannel (for example shown in Figure 1A, 4,5,13,14 and 17) accept to comprise in being suspended in biocompatible ferrofluid at least a target substance (for example, cell, bacterium, non-organism material etc.) sample, for example as described herein those.
After accepting sample, 1020 at least a electric currents (are for example controllably applied, use controller, for example based on the controller of processor, manually control etc.) at least two electrodes of contiguous microchannel to position (being even perhaps unitary electrode),, to produce controlled magnetic force in containing the ferrofluid passage of sample, make described at least a target substance guiding detecting device.Electrode can be for example any electrode configuration of Fig. 1-18 that relates to described herein.In some embodiments, detecting device can be based on capacitance measurement, or comprises the detecting device of other character (for example, impedance) of the sample of target substance based on measurement.This detecting device can capture region (for example, be configured to sample in the blanket of specific antibodies functionalization of the specific objective matter interaction that exists) the input and output stage determine the number (or amount) of target substance.Can show in the difference of the target substance number of input and output phase measuring the number that is configured the target substance member (for example, single organism) that (for example, functionalized) catch for the capture region of catching the specific objective material type.Therefore, based on the measurement that detecting device carries out sample in microchannel, at least a target substance in sample is determined 1030.
With reference to Figure 20, show the schematic diagram of general-purpose computing system 1100, it can be used for implementing any system/device based on processor, can be used in conjunction with system as herein described and equipment, comprises the controller 160 of Figure 1A, the evaluation unit 940 of Figure 17 etc.This computing system 1100 comprises the device 1110 based on processor, for example personal computer, special computing equipment, etc., it typically comprises central processor unit 1112.Except CPU1112, system comprises primary memory, cache memory, bus interface circuit (not shown).Device 1110 based on processor comprises large capacity storage element 1114, the hard disk drive that for example with computer system, is associated.Computing system 1100 may further include keyboard, or keypad 1116 and display 1120, for example, and the CRT(cathode-ray tube (CRT)) or the LCD(liquid crystal display) display.
Device 1110 based on processor is configured to be conducive to, for example, control as herein described and processing operation, for example, determine and/or control at least a controlled electric current that is applied to electrode, to produce controlled magnetic field, based on the character that detects (for example, electric capacity, impedance) determine the number of target substance/amount, etc.Thereby memory device 1114 also can comprise computer program, causes operating based on the device of processor when carrying out on the device 1110 based on processor, is beneficial to implement control as herein described and/or processes operation.Device based on processor can also comprise the peripheral unit that can exercise input/output function.These peripheral units can comprise, for example, CD-ROM drive and/or flash drive, or network connects, and is used for downloading related content to the system that connects.These peripheral units can also be used for downloading the software that comprises computer instruction, make system/device separately can carry out general operation.Alternatively and/or additionally, in some embodiments, the logical circuit of special purpose, for example, FPGA(field programmable gate array) or ASIC(application specific integrated circuit) can be used for the enforcement of system 1100.With described device based on processor 1110 can by together with other module of comprising be loudspeaker, sound card, pointing device, for example mouse or trace ball, the user utilizes them to provide and is input to computing system 1100.Device 1110 based on processor can comprise operating system, for example: the operating system of Windows XP Microsoft.Alternatively, can use other operating system.
In addition, various illustrative components, blocks, module and the method that relates to described in embodiment disclosed herein can use the general processor, digital signal processor (" DSP ") or other programmable logic device, discrete gate or transistor logic, discrete hardware components or its any combination that are designed to carry out function described herein implement or carry out.General processor can be microprocessor, but in replacement scheme, processor can be any processor, controller, microcontroller or state machine.Processor can also be implemented as the combination of calculation element, for example, and the combination of the combination of DSP and microprocessor, multi-microprocessor, one or more microprocessor and DSP kernel or any other this type of configuration.
Computer program (also referred to as program, software, software application or code) comprises the machine instruction for programmable processor, and can be with high-level program and/or OO programming language, and/or with assembling/machine language, implements.As used herein, term " machine readable media " (for example refers to any non-provisional computer program, equipment and/or device, disk, CD, storer, programmable logic device (PLD) (PLD)), be used for providing machine instruction and/or data to programmable processor, the machine readable media that comprises non-transience, it is accepted as the machine instruction of machine-readable signal.
For mutual with the user is provided, theme described herein can be implemented on computers, described computing machine (for example has display device, the CRT(cathode-ray tube (CRT)) or the LCD(liquid crystal display) display) be used for demonstration information to the user, and keyboard and pointing device are (for example, mouse or trace ball), utilize its user to provide and be input to computing machine.The device of other kinds also can be used to provide mutual with the user, and for example, the feedback that offers the user can be any type of sensory feedback (for example, visual feedback, audio feedback or tactile feedback); And user's input can be received in any form, comprises sound, voice or sense of touch input.
Partly or entirely can the comprising aft-end assembly (for example as data server) or comprise that middleware component (for example of theme as herein described, apps server) or comprise that front end assemblies (for example has graphic user interface or Web browser, by its user can be mutual with the embodiment of theme described herein client computer), or implement in the computing system of these rear ends, middleware or any combination of front end assemblies.The assembly of system can be by digital data communication (for example, the communication network) interconnection of any form or medium.The example of communication network comprises LAN (Local Area Network) (" LAN "), wide area network (" WAN ") and internet.
This computing system can comprise client and server.Client and server is generally all away from the other side, and it is interactive typically to pass through communication network.Relation between client and server usually relies on the computing machine that operates in separately, and has each other the computer program of client-server relation and produce.
Various illustrative components, blocks, module, circuit and relate to above-mentioned figure and embodiment disclosed herein described in method, often may be embodied as electronic hardware, computer software or both combinations.
In addition, relate to the method/program described in embodiment disclosed herein, can be embodied directly in hardware, the software module of being carried out by processor, or in both combinations.Software module can reside in the storage medium of RAM storer, flash memory, ROM storer, eprom memory, eeprom memory, register, hard disk, removable dish, CD-ROM or any other form, comprises in network storage medium.Exemplary storage medium can be coupled to processor, makes this processor can be from its reading information, and writes information in storage medium.In replacement scheme, this storage medium can be integrated into processor.Processor and storage medium also can reside in ASIC.
Some embodiments of the present invention have been described.Yet it will be understood to, can carry out various modifications and without departing from the spirit and scope of the present invention.Therefore, other embodiment is in the scope of following claim.
Annex A---the calculating of particle operation
Provided herein is to make particle speed and the threshold frequency of observing in ferromagnetic microfluid system described herein, apparatus and method can be by the analytical approach of evaluation., in order to simplify calculating, supposed the ideally ferrofluid of spherical incompressible particulate, magnet-wire, and be independent of the ratio of slip (seeing below) of magnetic field intensity and frequency.Further suppose the radius of spheric grain
(R)Compare the travelling-magnetic-field determined by electrode size and spacing (
H → ) wavelength be little, make
R|
H → |＜＜|
H → | set up.According to these hypothesis, the ferrofluid magnetization of next-door neighbour's particulate---with the volume of particle
VpThe interior effective magnetization
M eff ---it is uniform can being similar to.Also approximate is that any value (but not being its gradient) is constant at the Nei Yannei of particulate.
Then, the total transient force on this dipole by under establish an equation and provide
Wherein
B in The magnetic flux density in spherical particle, and to the granule interior volume integral (Zahn etc., 1995,
J of Magnetism and Magnetic Materials149:165-173).According to these hypothesis, the surface of equation A, due to
MWith
BUncontinuity to produce integration be 0 pressure term, the simple vector of integrand enlarges and shows, this is identical with Kelvin (Kelvin) force density.Therefore, the transient force expression formula can be reduced to
(equation B).
In order to obtain the analysis expression of final magnetic force, in the non-existent situation of particulate according to external magnetic field (
H ext ) represent
M eff And
B in Helpful, because this value can easily obtain from simple simulation.In ferrofluid (have complicated, the magnetic permeability of frequency dependence
μ p= μ 0 (1+χ
f )) have a magnetic permeability
μ P (be basically
μ 0 ) following depending on of net magnetization of particle
H ext
(equation C).
Determine that intragranular magnetic flux density and magnetic field need to consider the demagnetizing field that it is inner.Intragranular resultant field for spheroid is
H → in =
H → ext -
H → Dmag , with
H → Dmag =
M → eff / 3.Therefore, under linear condition, granulated magnetic can be written as
M → eff =χ
eff H → in , can obtain
(equation D).
Comparison equation C and D show the effective granulated magnetic rate aspect ferrofluid magnetic susceptibility:
Should be pointed out that effective magnetic susceptibility depends on ferrofluid, because particulate response magnetic force is only because it replaces ferrofluid, and generation " magnetic hole ".In this respect, the magnetic medium at place, hole determined the magnetic hole and apply the field between interactional intensity.Negative sign in equation E shows that the effective magnetization of particulate is the opposite direction of local ferrofluid magnetization under static condition.Although for χ
eff ≈ χ
f , for χ
f ＜＜1, the effective susceptibility in magnetic hole tends to-1 in the hard magnetization medium.In fact, use overgenerous ferrofluid to run counter to desire for particle manipulating.
Instantaneous magnetic force on particle can be expressed as
Here,
θBe
M → eff With
H → in Between angle, by resultant magnetization rate χ
eff Angle provide.Use
, we obtain
In the situation that there is row, the local magnetic field sinusoidal variations, and time average power be only transient force peaked half:
(equation H).
Similarly, the instantaneous torque on magnetic dipole by under establish an equation and provide
(Zahn?et?al.,?1995,?
J?of?Magnetism?and?Magnetic?Materials?149:165-173)。At this, the rotation of non-magnetic particles may be subject to the ratio of slip between 0 and 1
sThe possibility of impact be allowed to.This ratio of slip represents the ratio of the value of the torque that the isolated particle of non-magnetic particles torque and same size and effective magnetizing of experience in ferromagnetic microfluid system described herein and device will be experienced.
All the other detailed derivations and magnetic picture to magnetic moment are same.Replace magnetic flux density, obtain
Due to
, obtain
The time average torque is provided by following formula
For given input current amplitude, program of finite element (COMSOL) is used to calculate
H ext , use the realistic two-dimensional section of ferromagnetic microfluidic channel and lower electrodes.The Reynolds number (Reynolds'number) relevant with the motion of cell in static ferrofluid to the micro-meter scale pearl is very little.In this scheme, the impact of inertia can be ignored, stokesian fluid equation major flow body dynamics.Therefore, the balance between viscous drag and magnetic force determines granular dynamics.Because the stokesian fluid equation is linear, the fluid dynamics coefficient that all relate to can be incorporated in resistor matrix:
(Happel?J,?Brenner?1-1?(1983)?Low?Reynolds?Number?Hydrodynamics?with?special?applications?to?particulate?media.?(Martinus?Nijhoff:?Dordrecht))。Herein, ν is the linear velocity of particulate along passage length, and ω is its angular velocity, and η is ferrofluid viscosity,
RThe microballoon radius, and
f i Be depend on distance between particle radius and itself and channel roof (
h) the resistance factor.Suppose
H＜＜＜R,These resistance factors can obtain from the standard lubrication theory
(equation N)
(Goldman?et?al.,?1967,?
Chem?Eng?Sci?22:637-651)。Usually, likely by Dedaguin, Landau, Verwey and Overbeek theoretical (dlvo theory) (Ise, 2007,
Proc Jpn Acad B Phys. Mal Sci83; 192-198), use on particulate and the estimation of the surface charge density of channel surface
It, consider simultaneously the ion condition in ferrofluid.Ironically, particulate is pushed to the vertical direction of channel roof power (
F Avg, y ) be the magnitude of nN, and expect that they will be near the contact channels wall.
Equation M can ask ν and ω by a simple matrix inversion,
Wherein
Here,
G=f l f 4 -f 2 f 3 According to the facility on symbol, be defined.Therefore, because the particle linear velocity of magnetic force and torque can be determined separately:
Clean particle speed is provided by following formula
Magnetic force and magnetic moment particle volume (
R 3 ) weigh; , according to equation Q and R, be clear that the particle speed due to magnetic force depends on
R 2 , and because the particle speed of magnetic moment is weighed with R.This observations shows, the impact of magnetic moment on less particle is relatively more remarkable, and explained why less particulate they dynamically in demonstration less threshold frequency.
Aforesaid theoretical method well explain ratio of slip 1 and the about experimental result of the h of 1nm (for example, Figure 10 A), confirmed that particulate pushed to the expection of channel roof really consumingly.Ratio of slip 1 expression microballoon rotates under nonslipping condition.
Accessories B---the power on magnetic dipole
Usually, the magnetic force on magnetic dipole can use Kelvin's power expression formula to find, namely
(equation T).
This expression formula is about as much as the equation A that shows in appendix A.Crucial hypothesis is, the field that applies is not too inhomogeneous and particle radius (
R) enough little, make
R|
H → |＜＜|
H → | in any direction.According to this hypothesis, the magnetization that is right after particulate ferrofluid on every side can be thought uniformly.Further approximate is that any value (but not being its gradient) is constant at the Nei Yannei of particulate.
Remember the hypothesis of these simplification, the integral function that vectorial can be used for rewriting equation A is as follows:
First of equation U right-hand side (RHS) relates to the curling of magnetic flux density, can expand as
The curling of magnetic field of any position is 0 in the integration volume of equation A because ferrofluid and plastic particles be insulation and do not support electric current.Therefore, first of the RHS of equation V disappearance.The curling of particulate internal magnetization is 0, but on its surface, effectively magnetization changes as a step.Therefore, should usually be considered to the surface contribution of force density.Yet, because the magnetization that is right after particulate ferrofluid on every side is assumed that it is constant, when integration around spheroid, second also disappear (due to the symmetry) of equation V.Same logic can be applied to the B in equation U
→ x (
x M
→ ): microballoon inside (
x M
→ ) be 0, and B around on the ball surface
→ x (
x M
→ ) integration is 0, and is constant in magnetic flux density and the ferrofluid magnetization supposition at next-door neighbour microballoon place.
Based on same reason, (the B in equation U
→ 
) M
→ Item will be also 0 in microballoon inside, and integration is 0 around it.Unique item that relates to a non-zero gradient of vector is
Volume integral to non-magnetic particles is effective.Therefore, inner at particulate
B → =
μ 0 H → , and equation W becomes identical with equation T.In other words, according to the hypothesis of above emphasizing, equation A and T are in the situation that setting shown in this article is of equal value.
Equation A can be used as the expression formula of power, rather than in equation T, because the former causes power, its direction determined by gradient operator, need to take single derivative along given direction in space to determine the power along this direction.
What also consider is that what has occurred surface graded that in equation A, expression formula is correlated with.Again, using the magnetization of ferrofluid and magnetic flux density around microballoon inside and surface (but not being their derivant) is this constant hypothesis, and it can be defined as
Here, field and magnetization vector are on the x-z plane, due to the symmetry of ferromagnetic microchannel.As previously mentioned, the expression in equation X is only estimated in particulate inside, to calculate the power of x direction.If there is no general any loss, the center of microballoon is taken as initial point.
M → With
B → Be discontinuous on particle-ferrofluid border, so their derivant cause pulse, when at the microsphere surface integration,
M X, out B X, out – M Z, in B Z, in To from x=√ (
R 2–
y 2–
z 2) the surperficial small pieces integration located each contribution by x=-√ (
R 2–
y 2–
z 2) the negative contribution of locating relative small pieces offsets.Result surface integration is 0.Due to the spheroid symmetry, right
F z In be also like this.Therefore,, according to described hypothesis, in particulate inside, equation A evaluation is obtained magnetic force on it.
Claims (55)
1. the device of the plurality of target material of separate out suspended in biocompatible ferrofluid, described device comprises:
Microchannel, comprise at least one sample inlet and at least one outlet, this microchannel has the passage length that extends between at least one sample inlet and at least one outlet, this microchannel is configured to accept basically continuous sample flow from least one sample inlet, this passage is configured to make sample to flow at least one outlet along passage length, and described sample comprises plurality of target material and biocompatible ferrofluid; With
A plurality of electrodes of the contiguous microchannel in position, a plurality of electrodes are configured to when electric current is applied to a plurality of electrode to produce magnetic field model along at least a portion passage length of microchannel, magnetic field model be configured to cause when at least a portion of microchannel is advanced when sample flow at least two kinds of plurality of target material in sample flow separated.
2. the device of claim 1, it further comprises power supply, and described power configuration is for controllably to apply electric current controllably to produce magnetic field model to a plurality of electrodes.
3. the device of claim 2, wherein said power configuration is to apply one or more the electric current with selected amplitude, institute's selected frequency and/or selected phase place, and wherein at least two of sample kinds of target substances is separated to small part based on one or more of selected amplitude, institute's selected frequency and the selected phase place of electric current.
4. the device of claim 1, wherein said a plurality of electrodes are configured to produce controlled magnetic force component and controlled magnetic moment component.
5. the device of claim 1, wherein said a plurality of electrodes be configured to cause at least two kinds of target substances with microchannel in basically separate on the direction of the direction perpendicular of continuous sample flow.
6. the device of claim 1, it further comprises stream generation unit, this stream generation unit is configured to produce basically continuous stream, described stream generation unit comprises following at least a: forcing pump, syringe pump, peristaltic pump, vacuum plant, can be by the structure of gravity flowage, and produce the device of capillary force.
7. the device of claim 1, wherein said sample comprises the material based on cell.
8. the device of claim 1, wherein said biocompatible ferrofluid comprise appropriate ionic species and promote the continuation of cell to control osmotic pressure on cell.
9. the device of claim 8, wherein said biocompatible ferrofluid comprises the approximately citric acid salt concentration of 5-200mM.
10. the device of claim 9, wherein said biocompatible ferrofluid comprises the approximately citric acid salt concentration of 40mM.
11. the device of claim 9, wherein said biocompatible ferrofluid comprises the approximately ionic strength of the design of 150mM, so that biocompatible ferrofluid is that grade is oozed and is fit to keep eukaryotic alive.
12. the device of claim 1, wherein said biocompatible ferrofluid has approximately 7.4 pH.
13. the device of claim 1, wherein said at least two kinds of target substance based target sizes are separated.
14. the device of claim 1, wherein said at least two kinds of target substance based target shapes are separated.
15. the device of claim 1, wherein said at least two kinds of target substance based target elasticity are separated.
16. the device of claim 1, wherein said at least two kinds of target substance based target forms are separated.
17. the device of claim 1, wherein said at least two kinds of target substances be based on the frequency of electrode separation, the electric current that applies, and one or more of the phase place of the electric current that applies are hunted down.
18. separate the method for plurality of target material, described method comprises:
Accept the continuous sample flow that comprises multiple target substance in being suspended in biocompatible ferrofluid basically at the entrance of microchannel;
Sample is passed through along microchannel; With
A plurality of electrodes to the position adjacent channel apply at least a controlled electric current, and described current arrangements is for along at least a portion passage length of microchannel, controllably producing magnetic field model, and are separated to cause at least two kinds of plurality of target material in sample.
19. concentrate the device that is suspended at least a target substance in biocompatible ferrofluid, described device comprises:
Microchannel, described microchannel is configured to accept to comprise the sample of at least a target substance and biocompatible ferrofluid, and at least a target substance in the sample of acceptance is concentrated in the inlet flow with relevant input width substantially; With
At least two electrodes of the contiguous microchannel in position, described at least two electrodes are configured at least when controlled electric current is applied to described two electrodes, produce controlled magnetic force in containing the sample of ferrofluid, the controllable magnetic that produces causes at least a target substance to be concentrated in the stream that the width that obtains is narrower than the relevant input width of inlet flow.
20. the device of claim 19, wherein at least partly according to the physical attribute of at least two electrodes, at least two electrodes of the contiguous microchannel in described position are configured to conduct the electric current that controllably provides and produce controlled magnetic force.
21. the device of claim 20, wherein said at least two electrodes comprise having following at least a structure: the substantially parallel layout of one or more corrugated shape, at least two electrodes basically of one or more straight shapes basically of at least two electrodes, at least two electrodes, at least two electrodes are the location of taper basically, and wherein said at least two electrodes are near each other gradually.
22. the device of claim 20, wherein said at least two electrodes be configured to cause at least a target substance flow into above and space in microchannel between at least two electrodes.
23. the device of claim 19, wherein said at least two electrodes are configured to produce at least two magnetic waves of backward-travelling wave field, to cause at least a target substance, focus between the magnetic wave of at least two generations near the center, border.
24. the device of claim 23, wherein said at least two electrodes comprise the electrode of a plurality of substantially parallel layouts.
25. the device of claim 19, wherein said at least two electrodes comprise the array of electrode, the electrode that wherein at least some the first electrod-arrays are relatively adjacent tapered location is basically arranged, so that the first electrode is configured to move closer to adjacent electrode, electrod-array is configured to produce magnetic force, with the stream that causes at least a target substance that produces up and adjacent electrode between form.
26. the device of claim 19, wherein said at least two electrodes are configured at least when the controlled electric current with dependent phase is applied to two electrodes, produce controlled magnetic force in sample, wherein the dependent phase of at least a electric current is different from the dependent phase of another electric current.
27. the device of claim 19, wherein said at least a target substance comprises at least two kinds of target substances, and wherein said a plurality of electrode be further configured to cause at least two kinds of plurality of target material in the concentrated stream that obtains separated.
28. the device of claim 19, wherein said at least two interelectrode spacings increase gradually.
29. the device of claim 19, wherein said at least two interelectrode spacings reduce gradually.
30. the device of claim 19, wherein said at least two arrangement of electrodes are at least one electrode layer.
31. the device of claim 30, wherein said at least two arrangement of electrodes are in a plurality of electrode layers.
32. the device of claim 19, wherein microchannel length is with the angle of about 0-90 degree at least a portion through at least two contiguous electrodes.
33. the device of claim 19, wherein said at least a target substance comprises the target substance based on cell.
34. concentrate the method for at least a target substance in microchannel, described method comprises:
Acceptance comprises the sample that is suspended at least a target substance in biocompatible ferrofluid, and at least a target substance in the sample of acceptance is concentrated in the inlet flow with relevant input width substantially; With
At least two electrodes of contiguous microfluid controllably apply at least a electric current to position, to produce controlled magnetic force in comprising the microchannel of sample,
Wherein magnetic force causes at least a target substance to focus in the stream that produces that width is narrower than the relevant input width of inlet flow.
35. the device of at least a target substance in test sample, described device comprises:
Microchannel, it is configured to accept the sample of the biocompatible ferrofluid that comprises at least a target substance and wherein suspend at least a target substance;
Determine the detecting device of at least a target substance in sample; With
At least two electrodes of the contiguous microchannel in position, described at least two electrodes are configured at least when controlled at least a electric current is applied to two electrodes, produce controlled magnetic force in comprising the sample of ferrofluid, the controlled magnetic force that produces causes at least a target substance guiding detecting device.
36. the device of claim 35, wherein at least partly according to the physical attribute of at least two electrodes, at least two electrodes of the contiguous microchannel in described position are configured to conduct at least a electric current that controllably provides and produce controlled magnetic force.
37. the device of claim 36, the physical attribute of wherein said at least two electrodes comprises the electrode structure of at least two electrodes, produce controlled magnetic force according to this attribute, the structure of described at least two electrodes comprises following one or more: the substantially parallel layout of one or more corrugated shape, at least two electrodes basically of one or more straight shapes basically of at least two electrodes, at least two electrodes, at least two electrodes are the location of taper basically, and wherein said at least two electrodes are near each other gradually.
38. the device of claim 35, wherein said at least two electrodes comprise electrod-array, and at least some of array the first arrangement of electrodes is the tapered location of relative adjacent electrode, make described the first electrode be configured to move closer to adjacent electrode.
39. the device of claim 35, wherein said at least two electrodes are configured at least when the controlled electric current with dependent phase is applied to two electrodes, produce controlled magnetic force in containing the sample of ferrofluid, the dependent phase of at least a electric current is different from the dependent phase of another electric current.
40. the device of claim 35, wherein said sample comprises the plurality of target material, and wherein device further comprises:
The electrode group, be configured to when controlled electric current is applied to this electrode group, along at least a portion of microchannel length, produces controlled magnetic field model, cause at least two kinds of plurality of target material in sample separated.
41. the device of claim 35, wherein microchannel is configured to accept the sample flow of from external sample, originating.
42. the device of claim 35, wherein said at least two interelectrode spacings increase gradually.
43. the device of claim 35, wherein said at least two interelectrode spacings reduce gradually.
44. the device of claim 35, wherein said at least two arrangement of electrodes are at least one electrode layer.
45. the device of claim 35, the wherein length of the microchannel at least a portion of with the about angle of 0-90 degree, crossing at least two electrodes.
46. the device of claim 35, wherein said at least a target substance comprises the target substance based on cell.
47. the device of claim 35, wherein said detecting device comprises:
The electrode pair at interval, to measure the electric capacity in microchannel; With
Existing of at least a target substance determined with the electric capacity based on measured in the evaluation unit.
48. the device of claim 47, wherein saidly be configured to determine that at least a target substance exists identifies that cell location is the existence that at least a target substance is determined in the variation of the electric capacity in the measured microchannel that causes based on the existence due at least a target substance.
49. the device of claim 47, wherein said detecting device further comprises:
Capture region, it comprises and being configured to and the interactional material of one of plurality of target material, described capture region is positioned at the microchannel downstream of the electrode pair at interval; With
Another electrode to the interval that is positioned at the capture region downstream, to measure the electric capacity in microchannel;
Wherein saidly identify that cell location is to determine the initial number of at least a target substance in the electrode pair at interval based on the electric capacity that the electrode pair at interval is measured, determine the end number of at least a target substance with the electric capacity based at another place of electrode to interval, measuring, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
50. the device of claim 35, wherein said detecting device comprises:
The test set that the order of series connection arranges, the test set of the order setting of each series connection comprises:
The electrode at first pair of interval, to measure the electric capacity in microchannel,
Capture region, it comprises and being configured to and the interactional material of one of plurality of target material, described capture region is positioned at the downstream of the electrode at first pair of interval, and
Second pair of electrode that is positioned at the interval in capture region downstream, to measure the electric capacity in microchannel; With
Identify unit, determine the initial number of at least a target substance at the electrode place at first pair of interval with the electric capacity of measuring based on the electrode place at first pair of interval at each test set place, determine the end number of at least a target substance at the electrode place at second pair of interval with the electric capacity that electrode place based at second pair of interval is measured, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
51. the device of claim 35, wherein said detecting device comprises:
The electrode pair at interval, to measure the impedance in microchannel; With
Existing of at least a target substance determined with the impedance based on measured in the evaluation unit.
52. the device of claim 51, wherein saidly be configured to determine that at least a target substance exists identifies that cell location is the existence that at least a target substance is determined in the variation of the impedance in the measured microchannel that causes based on the existence due at least a target substance.
53. the device of claim 51, wherein said detecting device further comprises:
Capture region, it comprises and being configured to and the interactional material of one of plurality of target material, described capture region is positioned at the microchannel downstream of the electrode pair at interval; With
Another electrode to the interval that is positioned at the capture region downstream, to measure the impedance in microchannel;
Wherein saidly identify that cell location is to determine the initial number of at least a target substance in the electrode pair at interval based on the impedance that the electrode pair at interval is measured, determine the end number of at least a target substance at another place of electrode to interval with the impedance based at another place of electrode to interval, measuring, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
54. the device of claim 35, wherein said detecting device comprises:
The test set that the order of series connection arranges, the test set of the order setting of each series connection comprises:
The electrode at first pair of interval, to measure the impedance in microchannel,
Capture region, it comprises and being configured to and the interactional material of one of plurality of target material, described capture region is positioned at the downstream of the electrode at first pair of interval, and
Second pair of electrode that is positioned at the interval in capture region downstream, to measure the impedance in microchannel; With
Identify unit, determine the initial number of at least a target substance at the electrode place at first pair of interval with the impedance of at each test set place, based on the electrode place at first pair of interval, measuring, determine the end number of at least a target substance at the electrode place at second pair of interval with the impedance that electrode place based at second pair of interval is measured, and whether exceed predetermined threshold value based on determine the to be hunted down level of at least a target substance that zone catches of the difference between initial and end number at least partly.
55. the method for at least a target substance in test sample, described method comprises:
Accept to comprise the sample of at least a target substance in being suspended in biocompatible ferrofluid at microchannel;
At least two electrodes of contiguous microchannel controllably apply at least a electric current to position,, to produce controlled magnetic force in comprising the ferrofluid passage of sample, to cause at least a target substance, are directed to detecting device; With
At least a target substance in sample is determined in the measurement of sample in microchannel being carried out based on detecting device.
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| PCT/US2011/039516 WO2012057878A1 (en) | 2010-10-28 | 2011-06-07 | Microfluidic processing of target species in ferrofluids |
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| CN104535783A (en) | 2015-04-22 |
| US20130313113A1 (en) | 2013-11-28 |
| CN104535783B (en) | 2017-05-24 |
| EP2633330B1 (en) | 2022-05-11 |
| EP2633330A1 (en) | 2013-09-04 |
| WO2012057878A1 (en) | 2012-05-03 |
| EP2633330A4 (en) | 2018-01-10 |
| CN103403557B (en) | 2014-12-24 |
| EP4105656A1 (en) | 2022-12-21 |
| US9999855B2 (en) | 2018-06-19 |
| JP5809282B2 (en) | 2015-11-10 |
| AU2011320908A1 (en) | 2013-05-23 |
| JP2014500949A (en) | 2014-01-16 |
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